Micropara Midterm Coverage PDF

LESSON 1: THE MICROBIAL WORLD STRUCTURE & ACTIVITIES OF MICROBIAL CELLS Microbial cells Lecture Outline:...

LESSON 1: THE MICROBIAL WORLD STRUCTURE & ACTIVITIES OF MICROBIAL CELLS Microbial cells Lecture Outline: - living compartments that interact with their environment and with other - Microorganisms cells in dynamic ways - Structure and Activities of Microbial Cells - Microscopy and the Origins of Microbiology - The Impact of Microorganisms on Human Society MICROORGANISMS - Also called microbes - Life forms too small to be seen by the naked eye - Diverse in form and function - (inhabit every env that supports life) - (many of these microbes are undifferentiated single-cell organisms) - Live in microbial communities - Their activities are regulated by interactions with each other, with their environments, and with their organisms - (two fundamental types of cells: prokaryotic & eukaryotic) - (Difference between the two: - Size: smaller prokaryotic - Structure: prokaryotic simpler - EU - has membrane-bounded organelles; PR none - EU means true (has true nucleus) - genetic material is usually in the form of DNA found in nucleus and the nucleus is membrane-bounded - PR has NO true nucleus; has nucleus but not membrane-bounded; nucleus can be found in nucleoid - (Similarities) - Has cytoplasmic membrane - it is a permeability barrier - gatekeeper - resp. For substances that go and out of the cell - separates in and outside of the cell - Cytoplasm is the inside of the cell - mixture of macromolecules - has small organic molecules - various organic ions and ribosomes inside cytoplasm - PR and EU both have plasma membrane and cytoplasm - PR and EU have ribosomes - responsible for protein synthesis - PR and EU have genetic material in the form of DNA - In some cases, EU have cell wall but not all RED (ALL CELLS) - Metabolism - Cells take up nutrients. They transport and expel waste materials - w/c is why virus is not considered as living entity/things because it cannot metabolize. It needs a host. It is an obligate parasite - Growth - They can take nutrients from the env. And convert it to new cell materials to form new cells  - Evolution A BRIEF HISTORY OF LIFE ON EARTH - PR is faster due to their size - Viruses can also evolve but cannot do on its own BLUE (SOME) - Differentiation - Can form new cell structures such as spore (certain genera of bacteria only) - Spore formation: env is devoid of nutrients; they become dormant - Communication - Communicate via chemical messenger - Genetic exchange Microbes - oldest form - evolve to perform critical functions to support life - Horizontal type that is common in bacteria - Motility Earth - 4.6 bya - Some has flagella; enables the cell to move Microbial cells - betw. 3.8 and 4.3 BYA Evolved as anoxygenic phototrophic bacteria - able to utilize the energy from the sun to manufacture their own food BUT do not produce oxygen Anoxic Earth - no oxygen has diff gases Oxygen appeared in cyanobacteria - able to undergo photosynthesis, manufacture own food, and undergo oxygenic photosynthesis - convert energy from the sun into usable form of energy and has production of oxygen From anoxic to oxygenated atmosphere called GREAT OXYGENATION EVENT Allowed for evolution of modern UE and algal diversity then we have shelly inv and vascular plants then mammals then humans DOMAINS OF LIFE: BAE - Bacteria, Archaea, and Eukarya Light microscope B A - PRO; E - EU B - 1 kingdom A-1 EU - 4 (Plantae, animalia, protist, and fungi) then divided to phila Origin of their evolution -> LUCA (Last Universal Common Ancestor) MICROSCOPY & THE ORIGINS OF MICROBIOLOGY Robert Hooke - light is focused on the specimen by a glass condenser lens - English mathematician - image is then magnified by an objective lens and an ocular lens for - Coined the term “cell” projection on the eye - Observed cork under microscope - (Visible light is used pass through the specimen then lenses retract the - Authored “Micrographia” devoted to microscopic images light, then image will be magnified) - PARAMETERS IN MICROBIOLOGY Van Leeuwenhoek - Magnification - First to see bacteria under the microscope - Ratio of object, image to its real size - Constructed simple microscope - Resolution - Observed human blood smear - Measure of clarity of image - Contrast Electron Microscope - Made due to resolution problem of light - Resolution is inversely related to wave of light - Beam of electron shorter = better resolution - beam of electrons is used instead of light - electron beam is focused on the specimen by a condenser lens - Coated with gold -  SCIENTIFIC DEVELOPMENT OF MICROBIOLOGY Louis Pasteur (1822 - 1895) - known as the “Father of Microbiology” (aspetic technique, vaccines) - disproved the theory of spontaneous generation *Spontaneous Generation (Abiogenesis) - living organisms could arise from nonliving matter - Started in time of Aristotle and persisted for several centuries SEM - detailed structure of topography (surface) TEM - internal structure of the cell Disadvantage of electron microscope - preparation could kill your cells Francesco Redi - made the setup above (Meat Maggot Exp) 1st open, 2nd sealed, 3rd covered with gauze John Needham - did not believe Redi; published his own; heat broth for minutes then seal it. In one droplet, microbes are alive Lazaro Spallanzani - contradicted Needham. Performed several experiments including Needham’s. The duration of boiling matters to kill microbes  The Paris Academy of Sciences offered a prize to solve the dilemma. 1st setup no maggots/contamination. There's a filter 2nd setup - broke the next and exposed it Edward Jenner (1749 - 1823) - known as the “Father of Immunology” - Vaccine for smallpox BRANCHES OF MICROBIOLOGY - BACTERIOLOGY: study of bacteria - MYCOLOGY: study of fungi John Lister (1827 - 1912) - PHYCOLOGY: study of photosynthetic eukaryotes - known as the “Father of Modern Surgery” - PROTOZOOLOGY: study of protozoa - applied Pasteur’s work and introduced antiseptic techniques in surgery - VIROLOGY: study of viruses Robert Koch (1843 - 1910) - known as the “Father of Bacteriology” THE IMPACT OF MICROORGANISMS ON HUMAN SOCIETY Microorganisms, agriculture, & human nutrition Microorganisms as agents of disease Legumes plants - has associated of rhizobium (nitrogen fixing bacteria) live in root nodules. They convert nitrogen to a usable nitrogenous compounds Legumes give home to these bacterias and food (carbohydrates and lipids) - 1900s leading is microbial diseases Can be used as sustainable agri technique - Today not anymore Animal nutrition - ruminants (cow) - they have 4 stomach chambers Rumen - microbial fermentation - w/c is why they can digest cellulose - produced patty acids and waste products CO2 and CH4 these are greenhouse gases that contribute to climate change  Has fermented products Microorganisms and Industry Probiotics help in digestion Lactobacillus good bacteria Biofuels Microorganisms and Food LESSON 2: MICROBIAL CELL STRUCTURE & FUNCTION Lesson Outline 3d cubes - sarcina; grape-like structure - staphylococcus - Cells of Bacteria and Archaea - The Cell Membrane and Wall Some morphologies are immediately recognizable - Cell Surface Structures and Inclusions Spirochete – tightly coiled - Cell Locomotion - Eukaryotic Microbial Cells Form extension of their cells as long tubes or stalks budding and appendaged form Cells of Bacteria and Archaea Filamentous long chains of cells What is morphology? - In microbiology, it means cell shape Many variations cell morphology form a continuum some forms are very common but has variations Less common – budding type, spirillum, filamentous Coccus - spherical Rod - cylindrical Morphology is incorporated in their names Spirillum - Curved loose spiral shape Some cells of bacteria remain in groups/cluster after cell division – 2 largest thiomargarita and epulopiscium characteristic cocci – long chains Vary shape, sizes, cell volume Altho it is easily determined (morpho), but poor predictor of properties of cells Surface-to-Volume Ratio (i.e., gram staining) (many rod shaped archaea are indistinguishable from rod shaped bacteria) Cannot determine if bacteria is pathogenic or not Pathogenic – cause disease If under what kingdom (ar or bac) KNOW THE MORPHOLOGY FIRST then find out more using other tests - Small cells have more surface area relative to cell volume than do large cells and thus have a higher surface-to-volume ratio. S/V Ratio - The higher S/V ratio of small cells supports a faster rate of nutrient and waste exchange per unit cell. - Because of their small sizes, prokaryotic cells can grow faster and evolve more rapidly than can larger cells. - (Small cells -> large surf area – higher s/v - Importance – controls property growth rate and evolution - Growth depends on metabolism. Mas mabilis for small cells to metabolize - Prokaryotic cells faster than larger cells - Bacterias adapt mechanisms) The Cell Membrane and Wall The Cytoplasmic Membrane - surrounds the cytoplasm and separates it from the environment - (Cell membrane – cytoplasm (mix of macro and micro) – sep cyto to ooutside and envi - Physically c.m. rather weak but ideal structure for major cellular function – selective permability – filters what enters and what leaves) - the cytoplasmic membrane controls at least three critically important cellular functions: o permeability barrier - prevent passive leakage of solutes into and out to the cell - Structure o protein anchor - anchors sev protein that catalyze cell functions - BAE – Bacteria, Archaea, Eukarya 93 domains of life) o energy conservation - bac and archaea plays a role in energy - B & A – Prokaryotes consumption and conservation - Eu – eukaryotic - Cytoplasmic membrane of all cells is made up of phospholipid bilayer – hydrophobic and hydrophilic – philic water lover – phobic water repelling - Head – philic - Phobic - tail - B & E – phobic component consists of fatty acids; philic made up of glycerol – gly has phosphate – has functional group – can be sugar, coline, ethanolamine – it makes up ur glycerol - Fatty acids and gly – bonded together w/ ester linkage - B & e same structure - Inner surf cell membrane – cytoplasm Archaeal Membranes Archaeal membrane may form a lipid bilayer, monolayer, or mixed Bacterial Cell Wall - protection against osmotic lysis - confer shape and rigidity on the cell - (Bacterial cell wall – cytoplasm of cell maintains high concentration of dissolved solutes – this create high osmotic pressure The ester linkage in Archaeal membrane is the ETHER linkage. Instead of - This pressure is the osmotic lysis too much pressure – this is the cell fatty acid, hydrophobic region is repeating units of hydrocarbon 5 carbon wall to prevent that from happening which is isoprene - Certain antibiotics target bacterial cell wall synthesis Differentiate archaeal and bacteria and eukarya - Penicillin - first antibiotic - target bacterial wall synthesis - no cell wall - - Has ether linkage (archaea) subjected to osmotic lysis - Ester linkage for b n e - Both has glycer – the tail is diff composition – b and e fatty acids hydrocarbons called isoprene in archaea - Crenarchaeol has rings of their side chains and tails; cell membrane is varied; - Diglycerol tetraethers - at one end, tail is sandwiched  Peptidoglycan Cell of bacteria can be divided into two major groups: gram + and gram Gram positive – purple/blue (usual one) Gram negative – pink/red consists of two layers Surface of gram pos and neg looks Structure of Peptidoglycan (Bacteria) - rigid polysaccharide - found in all Bacteria that contain a cell wall - not present in the cell walls of Archaea or Eukarya ( consisted of the cell wall. Not present in Ar and Euk. Good for confering structural Sum bacteria do not have cell wall It is composed of alternating repeats of 2 modified glucose residue: - N-Acetylglucosamine (G) - N-Acetylmuramic acid (M) Glycan tetrapeptide – basic unit of peptidoglycan. Long chain of G.T. -> Peptidoglycan G and M bonded together by glycosidic bond – connected bia beta14 linkage The Gram-positive Cell Wall – this linkage can be destroyed by lysozyme – it can be seen in human secretion – major line of defense against bacterial infection B14 can be destroyed by penicillin - many Gram + Bacteria form several layers of peptidoglycan stacked one upon another - produce acidic molecules called teichoic acid The image above is the peptidoglycan structure in E. coli and Staphylococcus - (Gram positive – produce acidic molecules – teichoic acid – embedded aureus on their cell wall – composed of glycerol phosphate or ribitol phosphate (sugar + phosphate) attached molecule as glucose and alanine E. Coli - negative - Individual molecules are connected thru phosphate group to produce Staphylococcus Posit teichoic acid long strands – covalently linked to peptidoglycan In Gram + , there is a glycene interbridge, series of glycan tetrapeptide – - Lipoteichoic acid – covalently bonded to membrane lipids than gram positive bacteria only peptidoglycan – functions to help in divalent metal ions (irons, magnesium ions) to bind divalent metal ions prior to their transfer to the cells. The Gram-negative Cell Wall O-specific polysaccharide - Typically contains galactose, glucose, rhamnose, and mannose, and one or more dideoxyhexoses Core polysaccharide - In Salmonella species, the core polysaccharide consists of - only small amount of the total cell wall consists of peptidoglycan ketodeoxyoctonate (KDO), various heptoses, glucose, galactose, and - most of the wall is composed of the outer membrane (LPS) Nacetylglucosamin - Thinner Lipid A - Lipid portion of LPS; include fatty acids such as: - Cell walls have outer membrane – effectively 2nd bilayer but not o caproic acid cytoplasmic membrane – composed of polysaccharide and lipids – o lauric acid linked together to form complex – outer membrane or LPS o myristic acid lipopolysaccharide layer o palmitic acid LPS - Acts an effective barrier against other harmful agents that would o stearic acid penetrate the plasma membrane - Toxicity is specifically linked to the LPS layer, in particular, to lipid A. - Endotoxin- toxic component of LPS. Knowledge on cell wall of microbes is important (some gram negative do (Polysaccharide portion of LPS: Core and O not work with antibio) Salmonella species, LPS is well studied O-> core connected to O-specific.; contains diff sugars o-specific has more contact outside Lipid A – fatty acids; Toxicity – important bio activity toxicity to animals bc it releases endotoxin Binding proteins – begin the process of trasnporting substraits Gram negats have endotoxin- if infected – render endotoxin – poision/toxic to Chemoreceptors – proteins that govern chemotaxis response cell Proteins that chuchu – can be find in preliplasm; galing sa precursor molecules secreted thru CM – this cause extracellular structures) The Periplasm and Porins Porins - Proteins that function as channels for the entrance and exit of solutes PORINS – Gram negats; Braun lipoprotein – molecules that spans the gap betw and LPS layer and peptidoglycan layer; like an anchor; Archaeal Cell Wall - variety of cell wall structures are found in Archaea, including walls containing polysaccharides, proteins, or glycoproteins or mixture of these macromolecules Very diverse; variety of cell wall structures found in Archaea Periplasm - May contain several classes of proteins o Hydrolytic enzymes o Binding proteins o Chemoreceptors o Proteins that construct extracellular structures (Periplasm – space loc between outer surface of CM and inner surface of outer membrane Hydrolytic enzyme – cyloritic enzdegrade polymeric substances  B13 linkage – insensitive to lisozyme and penicillin Amino acids are all of L stereoisomer Pseudomeurein & Other Polysaccharide Cell Walls Cells wall of some other Archaea lack pseudomurein and instead contain other polysaccharides i.e Methanosarcina species have thick polysaccharide walls composed of glucose, glycuronic acid, galactosamine uronic acid, and acetate S-Layer - paracystalline surface layer - most common type of cell wall in Archaea PSEUDOMUREIN - consist of interlocking molecules of protein or glycoprotein - backbone is formed by alternating repeats of N- acetylglucosamine - sufficiently strong to withstand osmotic pressures without any other and N- acetyltalosaminuronic acid wall components - glycosidic bonds between the sugar derivatives is -1,3 - the amino acids are all of L stereoisomer - immune from destruction by both lysozyme and penicillin Pseudomurein – like a peptidolgcan byt diff backbones by alternating G connected to T Polysacc but backbone by repeats of _______ Cell Surface Structures and Inclusions Slime Layer Cell Surface Structures - Capsules and Slime Layers o Capsules and slimes layer are often interchangeable o Tight mmatrix – capsule o Deformed – slime layer o Both attach bacteria to surface - Fimbriae, Pili, and Hami - layer that is more easily deformed, loosely attached, and does not Capsule exclude particles Fimbriae - layer organized in a tight matrix that excludes small particles and is - enable cells to stick to surfaces, in case of animal tissues in the case tightly attached of pathogenic bacteria, or to form pellicles (thin sheets of cells on a - readily visible by light microscope if cells are treated with India ink liquid surface) or biofilms on surfaces  - molecules that contribute to the pathogenicity of a bacterial pathogen Pili - i.e. the case of Bacillus anthracis Cell Inclusions - Carbon Storage - Polymers Polyphosphate, Sulfur, and Carbonate Minerals - Magnetic Storage Inclusions: Magnetosomes - Gas Vesicles - similar to fimbriae but are typically longer and only one or a few pili are - Endospores present on the surface of the cell CARBON STORAGE POLYMERS - facilitate genetic exchange between cells in the process called conjugation (conjugative or sex pili) - enable the adhesion of pathogens to specific host tissues that they subsequently invade (type IV and other pili; twitching motility (type IV) Hami - Poly- -hydroxyalkanoate (PHA) o synthesized by cells when there is an excess of carbon and are - unique attachment structure that resembles a tiny grappling hook broken down as carbon energy sources when conditions - structurally resemble type IV pili except for their barbed terminus, warrant which functions to attach cells both to surfaces and to each other o If there is an excess of carbon source – synthesize – store form - grappling hook – similar structure to type IV pili – only observed in of PHA – they can breakdown this if there's a scarcity Archaea - Glycogen o polymer of glucose; reservoir of both carbon and energy and is Virulence Factors produced when carbon is in excess  o excess carbon MAGNETIC STORAGE INCLUSIONS: MAGNETOSOMES - Magnetosomes POLYPHOSPHATE, SULFUR, AND CARBONATE MINERALS - Polyphosphate Granules o formed when phosphate is in excess and can be drawn as a o enable some bacteria orient themselves within a magnetic field source of phosphate for nucleic acid and phospholipid o biomineralized particles of of the magnetic iron oxides magnetite biosynthesis when phosphate is limiting and greigite o phosphate excess o Magnetotaxis: the process of migrating along Earth’s magnetic field - Sulfur Storage Products o oxidized reduced sulfur compounds GAS VESICLES o Can be used if sulfur is limuted - Gas Vesicles - Carbonate Minerals o structures that confer buoyancy and allow cells to position o Gloeomargarita: forms intracellular granules of bensonite themselves in regions of water column that best suit their metabolism  o (Gas vesicles – example is buoyant cyanobacteria – they need o sporulation occurs when a key nutrient becomes limiting sunlight – go to a water column best suited for their metabolism – if photosynthetic go to have sunlight) - Endospores o an endospore can remain dormant for years but can convert back to vegetative cell rapidly o three steps: activation, germination, and outgrowth ▪ activation: occurs when endospores are heated for o highly differentiated cells that are extremely resistant to heat, several minutes at an elevated but sublethal temperature and harsh chemicals ▪ germination: typically a rapid process; loss of refractility o function as survival structures and enable the organism to ▪ outgrowth: involves visible swelling endure unfavorable growth conditions o Survival structures - endosporulation Endospore – survival structures – endosporulation - Endospore Formation and Germination When this happen Vegegative cell – regular functioning cell can be converted to chuchu. This can be converted to something not to die. Ie., hibernate forming of endospores – maturw If an envi becomes nutrient limiting Can remain dormant for years but can be converted back to vegetative -> go to germination Refractile – refract light o a vegetative cell is converted into a non growing, heat- resistant, and light- refractive structure Bacillus anthracis hard to die in dormant/endo – can only be killed during vegetative - Structure of Bacterial Endospore Importance of Calcium and DPA – they form complex w/c is the calcium dipo chuchu acid – form 10 percent of dry weight of the endospore function to bind freewater w/n the endo helping to dehydrate the dev endo; DPA compk insert o exosporium: thin protein covering between basis and dna para mastabilize ang dna against heat degeneration o spore coats: layers of spore- specific proteins o cortex: loosely cross- linked peptidoglycan o core: contains core wall, cytoplasmic membrane, cytoplasm, nucleoid, ribosomes, and other cellular essentials Cell Locomotion Flagella Structure and Activity - Flagella, Archaella, and Swimming Motility - Gliding Motility Bacterial Flagella - sing. flagellum - long- thin appendages; free at one end and anchored into the cell at - filament: main part; composed of many copies of flagellin the other end - hook: connects filament to flagellum motor - Flagellin is protein - flagellum motor: has two main components: rotor and stator - rotor: central rod and the L, P, C, and MS rings - stator: consists of Mot proteins that surround the rotor and function to generate torque - rotation of flagellum occurs at the expense of the proton motive force Archaella - flagella can rotate up to 1000 revolutions per second to support a swimming speed of up to 60 cell lengths/ sec  Eukaryotic Microbial Cells - sing. archaellum - The Nucleus and Cell Division - impart movement to the cell by rotating - Mitochondria, Hydrogenosomes, and Chloroplasts - filament is made up of several different proteins - Other Eukaryotic Cell Structures - overall structure bears resemblance to type IV pili - rotation is driven by hydrolysis of ATP The Nucleus - Methanocaldococcus swims nearly 500 cell lengths per second, which makes it the fastest organism on Earth! - Thinner than flagella Gliding Motility - some bacteria are motile but lack flagella - these non swimming yet motile cells move by gliding - gliding bacteria are typically filamentous or rod- shaped in morphology - gliding process requires that the cells be in contact with a solid surface - contains the chromosomes of the eukaryotic cell - histones: proteins that tightly pack the DNA to form nucleosomes - nucleolus: site of rRNA synthesis Cell Division Chloroplasts - chlorophyll- containing organelles of phototrophic microbial eukaryotes that carry out photosynthesis - If photosynthesis occurs, there are chloroplasts - mitosis: unique to eukaryotic cells o during mitosis, chromosomes condense, divide, and are separated into two sets, one for each daughter cell - meiosis: converts a diploid cell into several haploid cells Mitochondria - site of respiration (aerobic eukaryotic cells) - enclosed by a double membrane system Hydrogenosomes - present in anaerobic eukaryotic microorganisms - lack citric acid cycle enzymes and cristae - metabolism is strictly fermentative - oxidation of pyruvate to hydrogen, carbon dioxide, and acetate LESSON 3: Microbial Taxonomy Taxonomy - the science of classification; - provides an orderly basis for the naming of organisms and for placing organisms into category, or taxon (plural: taxa) Microbial Taxonomy - the study of the diversity of microorganisms with the aim of organizing and prioritizing in an orderly manner. THREE COMPONENTS OF TAXONOMY 1. Identification - the process of characterizing organisms - (Morphology first, further tests biochemical tests) 2. Classification - the process of arranging organisms into similar or related groups We also use the polyphasic approach - take into consideration its molecular 3. Nomenclature basis aside from morphological basis - the system of assigning names to organisms Taxonomy reflects diversity of life Bergey’s Manual of Determinative Bacteriology (9th edition) - The accepted reference on the identification of bacteria is commonly referred to as the Bergey’s Manual of Determinative Bacteriology by David H. Bergey. Carolus Linnaeus - Swedish botanist credited with founding the science of taxonomy - Originated binomial nomenclature, “two name” system Binomial Nomenclature - Escherichia coli - Genus (plural: genera) + specific epithet - First letter of the genus is ALWAYS in uppercase; spec. epithet is ALWAYS in lowercase. They are ITALICIZED. If written manually, they are UNDERLINED  - time of Linnaeus - kingdom down to species; kingdom is the broad while species is the most specific USING A TAXONOMIC KEY - The name of an organism often tells something about it, such as its shape, where it is found, what nutrient it uses, who discovered it, or what disease it causes. Dichotomous Key - has paired statements describing characteristics of organisms - Linnaeus also established a hierarchy of taxonomic ranks - Presents an either or choice; one statement is true. Each statement is followed by directions to other pairs of statements - Domain pinakamalawak  Taxonomic Ranks - In taxonomic hierarchy, the lowest level is considered species. 1700s - Flora - plants - Fauna - animals 1866 (Haeckel) Added protista 1959 (Whittaker) - 5 Kingdom system 1990 - Introduced domains LESSON 4: Flow of Flow of Genetic Information: Genetic info is embedded in the seq of nucleotides in nucleic acids Replication, Transcription & Replication, Transcription & Translation in macromolecules. nucleic acids div in DNA and RNA Prokaryotes DNA carries blueprint RNA produced during transcription and it carries mRNA _. Carries the copy of PART 1: DNA REPLICATION blueprint of DNA - Templates and Enzymes + Steps in Replication mRNA - convert by rna to defined amino acid seqs in proeins - dna replicate - - The Origin of Replication & The Origin of Replication & The Replication one strand undergo trans to produce rna then translate to protein - protein Fork building blocks of organisms - collectively, nucleic acid + proteins - Bidirectional Replication Bidirectional Replication and the Replisome informational macromolecules REVIEW DNA RNA PROTEIN - Central Dogma of molecular biology Diff DNA and RNA - Diff in structure - RNA 1 strand DNA double strand - Pentose sugar RNA is ribose then DNA is deoxyribose - Nitrgoenous bases are diff (DNA AGTC) (RNA AGCU) Monomers of nucleic acids (monomer is the basic unit of molecule to do polymer; it is a larger molecules) - nucleotide - too many then polynucleutide Dna and RNA are polynucleotides THREE COMPONENTS NUCLEOTIDES Sugar - in the form of pentose - 5 carbon Phosphate molecules linked to sugar = sugar phosphate backbone Cells are chemical machines - transform nutrients into new cell materials, can Nucleotide differs based on their base - nitrogenous base (3rd components) be viewed as coding devices they use genetic information Purines - 2 guanine and adenine Gene - basic functional unit of gen info Pyrimidines - cytosine and thymine and uracil Genome - total complement of genetic elements; the totality  There is a bond which is the Hydrogen Bond (bond one nucleo from one strand to another) Dna molecules are arranged anti-parallel manner = 5’ and 3’; 3’ and 5’ BALIKTAD Wrapped around each other to form double helix Helix naturally forms two distinct grooves: major and minor grooves; difference is the space abundance. Most proteins that interacts with DNA bind with the major groove because of space abundance Nucleic acid backbone is a polymer of alternating sugar and phosphate molecules Sugar phosphate backbone (yung mukhang light green piattos) Between sugar & phosphate between nucleotides- the bond is called phosphodiester bond Sequence of nucleo in dna and rna molec ay the primary structure and encode genetic information To bond, there should be base pairing Erwin Chargaff’s observation that in the base composition of DNA, the A-T quantity of adenine equaled the quantity of thymine and the quantity of G-C guanine equaled the quantity of cytosine. A=T G=C DNA REPLICATION IN PROKARYOTES Double stranded molecule is copied to produce identical DNA molecule - DNA replication is semiconservative = each strand will contain one parent and one daughter - Each of the two progeny double helices have one parental and one new strand - Replication ALWAYS proceeds from the 5' end to the 3' end The precursor is deoxy - 1 nitrogenous base, sugar, and 3 phosphate  DNA and its enzyme add new nucleotide from the 5 prime end and 3 prime end direction (in leading strand); leading strand will always have 3 prime end The Origin of Replication and DNA polymerase III will add nucleo there and its enzyme is resp in adding complementary basis sa DNA strand. Two more important proteins: single strand binding protein (helps hold DNA in place during the replication process) and topoisomerase or DNA gyrase (helps ease the supercoiling of DNA strand; can be located beyond the replication fork) Before DNA polymerase III can add nucleo, it needs an RNA primer. The enzyme responsible for synthesizing this RNA Primer is called Primase. If RNA primer is there, DNA nucleotides can be added. It can only be added in 3 prime ends because it has 3-OH which new bases can be attached to. This is the reason why replication process always occurs from 5 prime to 3 prime Compare the bacterial genome from the eukaryotic genome direction - Bacterial genome has circular chromosomes - And smaller than eu 3’ has three hydroxyl group (it means it is easy for new nucleotides to attach - Lacking of introns (noncoding; exons are coding) full of exons to u) - Eukaryotes are full of exons and introns 5’ doesnt have this - DNA of EU is linear and bigger - DNA repli starts with OriC Leading strand - 5 to 3; replication is continuous. Why? It is easier to add new - 1 OriC for bacterial nucleotides due to three hydroxyl groups. DNA polymerase III will just add - Multiple OriC for Eu there Single defined sequence is about 245 base pairs in length = recognition site Lagging strand 3’ to 5’; not continuous replication; THAT'S the problem; discontinuous replication; it needs FRAGMENTS such as Okazaki fragments. DNA synth begins at OriC -> Helicase is an enzyme that when ready for DNA replication it will recognize OriC -> will unwind the DNA strand -. When Aside from DNAPIII, it needs another enzyme: DNA Polymerase I for the unwinded, replication bubble will form - separation of two strands will create V LAGGING strand. DNAP1 will remove RNA primer and replace it with DNA section called replication fork -> once opened, two strands will be visible - dna (Repli in lagging is by fragment. U need plenty of rna primers) once polymerase III will enter it is the primary enzyme in replicating chromosomal syntheized, dna polymerase III will take action. DNAP1 will remove primers with DNA. Since by fragments, thereare grapes. DNA ligase will connect and seal these fragments so that there will be a whole dna strand DNA has two strands: Leading Strand - Topoisomerase breaks and reforms DNA’s phosphate backbone ahead of the replication fork, thereby relieving the pressure that results from this supercoiling - Primase synthesizes an RNA primer with a free 3'-OH, which DNA polymerase I uses to synthesize the daughter strands. Lagging Strand - Occurs continuously on the leading strand Discontinuously on the DNA Gyrase - Topoisomerase lagging strand  Bidirectional Replication - Okazaki Fragments are on lagging strand - DNA synthesis is bidirectional in prokaryotes - Two replication forks moving in opposite directions  - REPLISOME: Complex of multiple proteins involved in replication. - DNA pulled through the replisome Tau protein holds replication enzyme together in the complex called replisome Repli is highly coordinated for both leading and lagging strand Proofreading in DNA - Proofreading occurs in prokaryotes, eukaryotes, and viral DNA replication systems - DNA replication is extremely accurate - Proofreading helps to ensure high fidelity - Polymerase can detect mismatch through incorrect hydrogen bonding - Mutation rates in cells are 10^-8 to 10^-11 errors per base inserted PART 2: RNA SYNTHESIS: TRANSCRIPTION RNA polymerase uses DNA DNA will be serving as the template to create an RNA strand—--- Only one strand wll be transcribed in the process of transcription. - Transcription 3 to 5 of lagging will be used = antisense strand - The Unit of Transcription Unlike DNA replication, both are needed. In transcription, only 1 will be - Transcription in Archaea and Eukarya transcribed/needed. TRANSCRIPTION - Transcription (DNA to RNA) is carried out by RNA polymerase - RNA polymerase uses DNA as template For trans to be initiated, DNA Pol should bind to a promoter region = site of - RNA precursors are ATP, GTP, CTP, and UTP initiation of transcription. DNA pol will stick to it. DNA Pol is the core enzyme - Chain growth is 5' to 3' just as in DNA replication in the trans process Transcription has 3 process - Initiation To find promoter, sigma factor will help DNA Pol - Elongation - Termination Transcription is basically the creation of RNA from the DNA molecule Genetic blueprint in DNA, RNA will copy it  Transcription: Termination - Termination of RNA synthesis is governed by a specific DNA Sigma factor will recognize two highly conserved regions of promoter: sequence; two types of termination Pribnow box (TATA box) and 35 region (consensus region) - Intrinsic terminators: Transcription is terminated without any additional factors (example of formation of loops; once loops are If the sigma factor sees the promoter site, it will guide DNA polymerase where formed, it will serve as roadblocks. Then it will signal the end of it should start. transcription) No need the for proteins help Elongation will start. Unlike DNA repli, where entire chromosomes are replicated, transcription only smaller portions are needed to be transcribed. - Rho-dependent Termination: Rho protein recognizes specific DNA sequences and causes a pause in the RNA polymerase For example, a certain protein is needed by the cell, then it will be the only (rho will bind a specific seq sa template and will terminate the one that will be transcribed. actions of DNA Pol) If the cell needs only gene 3, then it will be the only one that will be transcribed. No need to transcribe all. The Unit of Transcription One initiation site to one termination site is called one unit of transcription Bacterial cells - one operon - Unit of Transcription: unit of chromosome bounded by sites where transcription of DNA to RNA is initiated and terminated- - Most genes encode proteins, but some RNAs are not translated (i.e., rRNA, tRNA) - Three types of rRNA: 16s, 23s, and 5s - rRNA and tRNA are very stable - tRNA co-transcribed with rRNA or other tRNA - (some RNAs are noncoding) - Products of transc are RNA molecules. In these RNA molecules, there Polycistronic mRNA - mrna that encodes _______ will be those that code for proteins- these are mRNA - they will Prokar and eukar genomes undergo translation - they will be converted to as proteins Theres operon - NOn-coding RNAs are rRNA (ribosomal) and tRNA (transfer) 1 operon between initiation and termination there is 1 promoter region may - Prokaryotes have 70s ribosomes it has two parts - large (50s) and genes na magkakapareho small (30s) subunit - mRNAs have short half-lives (a few minutes) These similar genes are transcribed by one promoter called operon They are expressed multiple genes Transcription in Archaea and Eukarya Rare in archaea Eukrayotic has Exons and Introns which is why post-transcriptional modification happens DNA rep, transc, and translation, these three happens in cytoplasm of prokaryotic cells since prokar do not have organelles However, Eukar, DNA repli and transcript happen in the nucleus. Translation happens in cytoplasm because the EU has organelles. - The Archaea contain only a single RNA polymerase However, EU RNAs produced during transcription, have Introns. So, it is - Resembles eukaryotic polymerase II necessary to have post-transcriptional modification before it exits cytoplasm. Eukaryotic RNA processing happens through RNA splicing. Happens in the nucleus by removing introns in RNA transcripts before it exits the cytoplasms. Enzyme responsible is spliceosomes. - Eukaryotic RNA processing: many RNA molecules are altered before they carry out their role in the cell - RNA Splicing: When compared to gene regulation processes, archaea have more - Takes place in nucleus similarities w/ bacteria - Removes introns from RNA transcripts Archaea they are kind of the middle of eukarya and bacteria - Performed by the Spliceosome Introns cannot be found in bacteria  - Eukaryotic RNA processing -RNA capping - Addition of methylated guanine to 5' end of mRNA - Poly (A) tail - Addition of 100 - 200 adenylate residues - Stabilizes mRNA and is required for translation Then MATURE RNA EU - HAPPENS INSIDE THE NUCLEUS (DNA REPLI, AND TRANSCRIPTION) PRO - ALL IN CYTOPLASM IF RNA READY FOR TRANSLATION => PROTEIN SYNTHESIS PART 3: PROTEIN SYNTHESIS TRANSLATION - Polypeptides, Polypeptides, Amino Acids, Amino Acids, and the and the Peptide Bond - Translation and Translation and the Genetic the Genetic Code - Transfer DNA - Protein Protein Synthesis - Protein Folding Protein Folding and Secretion Building blocks of protein = amino acids that are linked together All amino acids = polypeptide = bonded by peptide bond = polypeptide then create protein PROTEIN Differences in the nature of amino acids differ in the R group (Serine, Threonine etc.) Translation and the Genetic Code Just like transcript - it has I,E,T Translation - synthesis of protein in RNA For RNA to be translated to a protein, certain bases to be read. These bases are called genetic code or codon. Codon is a triplet of nucleic acid bases. One codon = 1 amino acid - Proteins play a major in cell function Initiation = start codon = AUG = Methionine = (bacteria modified Methionione - Catalytic proteins (enzymes) called fMET w/c means N-Formylmethionine) - Structural proteins - Proteins are polymers of amino acids Termination = stop codon = UAA, UAG, & UGA - Amino acids are linked by peptide bonds to form a polypeptide 5’-AUGCCCGGGUUUUAC-3’ (RNA) Need genetic code to encode specific amino acid; triplets of nucleic acid bases Triplet 5’-AUG|CCC|GGG|UUU|UAC-3’ Each codon encode for specific amino acids Genetic code is a degenerate code there are multiple codons that encode a single amino acid Anticodon- tRNA; crucial in translation process; brings amino acids; anticodon carries anticodon Ensures that it brings the proper amino acid Last base is the wobble position; due to the degenerate feature of the genetic codes Wobble position would make the base pairing flexible - Translation: the synthesis of proteins from RNA - Genetic code: a triplet of nucleic acid bases (codon) encodes a single amino acid - Specific codons for starting and stopping translation - Degenerate code: multiple codons encode a single amino acid - Anticodon on the tRNA recognizes codon Start codon = initiate translation = elongation = terminate if encountered stop codon Proper reading frame Shine-Dalgarno sequence - reading frame Amino acid attached to phe Transfer RNA - Transfer RNA: at least one tRNA per amino acid - Bacterial cells have 60 different tRNAs - Mammalian cells have 100-110 different tRNAs - Specific for both a codon and its cognate amino acid - tRNA and amino acid brought together by aminoacyl-tRNA synthetases - ATP is required to attach amino acid to tRNA - tRNA is cloverleaf in shape Protein Synthesis - Ribosomes: sites of protein synthesis = this is where translation happens - Thousand of ribosomes per cell - Composed of two subunits (30S and 50S in prokaryotes) - S = Svedberg units - Combination of rRNA and protein - E. coli has 52 distinct ribosomal proteins Once polypeptide is formed, it is not automatically functional. It has to be folded. PROTEIN HAS TO BE FOLDED Protein Folding and Secretion - Most polypeptides fold spontaneously into their active form - Denaturation - Some require assistance from molecular chaperones or chaperonins - Occurs when proteins are exposed to extremes of heat, pH, or for folding to occur certain chemicals - They only assist in the folding; they are not incorporated into protein - Causes the polypeptide chain to unfold Can also aid in refolding partially denatured proteins - Destroys the secondary, tertiary, and/or quaternary structure of - (they only assist mole chap to fold. They do not become part of the protein protein) - The biological properties of a protein are usually lost when it is denatured Some proteins need to be secreted outside the cell (where they are needed) Needed by proteins that have to be folded inside so that once they go out, These proteins should not be folded before they go out of cells they can function immediately Example, structural proteins needed for plasma membranes. They will fold = go out tat protein export system = toxins example outside If they fold, it will be hard for them to go out of the cell Signal sequence = recognized by another protein signal recognition particle = this bind to signal seq = prevents protein to fold = easier to go out = exits in sec system and it will fold there