nucleic lec 11

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Questions and Answers

What aspect of TAL proteins makes them easy to engineer?

They possess a high affinity for RNA.

They can recognize any arbitrary DNA sequence.

Only residues 12 and 13 specify the base. (correct)

They require complex folding to function.

Why is the major groove more informative for DNA recognition by proteins?

It has a more varied pattern of hydrogen bonding groups. (correct)

It contains fewer hydrogen bond donors and acceptors.

Proteins bind more easily in the minor groove.

Its width allows for better protein binding.

What common structural feature do DNA binding proteins use to interact with bases?

a-helices inserted into the minor groove.

Hydrogen bonds to the edges of bases. (correct)

Hydrophobic interactions with the DNA backbone.

Beta sheets that span across entire nucleotide sites.

What role does the phosphate backbone play in DNA binding proteins?

Most of the binding energy comes from interactions with it. Signup and view all the answers

How do DNA binding proteins typically distort DNA?

By shifting it into an A-DNA like conformation. Signup and view all the answers

What does two-fold symmetry in DNA binding sites help achieve?

It expands the binding footprint on the DNA. Signup and view all the answers

What is the main reason that Nucleic Acid binding proteins undergo order-disorder transitions?

To facilitate the binding of proteins to nucleic acids. Signup and view all the answers

In the context of DNA binding, what role do hydrophobic interactions typically serve?

They contribute to the specific recognition of bases. Signup and view all the answers

What is the primary structure of nucleic acids composed of?

A sequence of deoxynucleotides Signup and view all the answers

Which of the following interactions is NOT typically present in the DNA backbone?

Hydrogen bonding Signup and view all the answers

What structure do the ribose-phosphate backbones of nucleic acids exhibit?

Multiple rotatable bonds allowing flexibility Signup and view all the answers

What type of biological molecule typically interacts with the regulatory methylation of bases in DNA?

Proteins Signup and view all the answers

Which of the following best describes the 5’ phosphate group in DNA?

It generally has a single negative charge. Signup and view all the answers

Which of the following statements concerning purines and pyrimidines is accurate?

Purines have two nitrogen atoms per ring, while pyrimidines have one. Signup and view all the answers

What defines the directionality of a nucleic acid strand?

The orientation of phosphate groups Signup and view all the answers

What role does ribose play in RNA compared to deoxyribose in DNA?

Ribose can act as an additional hydrogen bond donor. Signup and view all the answers

In the structural hierarchy of nucleic acids, what do the secondary structures primarily consist of?

Double stranded helices Signup and view all the answers

What stabilizes the structure of DNA more effectively than A-DNA?

Presence of a water spine in the minor groove Signup and view all the answers

Which of the following statements is true regarding Z-DNA?

Guanine tetrads can influence transcription. Signup and view all the answers

What is the main destabilizing factor for nucleic acid structures?

Electrostatic repulsion of phosphate groups Signup and view all the answers

Why does RNA usually lack a duplex formation like DNA?

Most RNA is transcribed from only one DNA strand. Signup and view all the answers

Which of the following contributes to the enthalpic stabilization of DNA duplex formation?

Hydrophobic interactions and base stacking Signup and view all the answers

What characteristic of A-DNA differentiates it from B-DNA?

Presence of a hole down the helix axis Signup and view all the answers

What is a key feature of the major groove in B-DNA?

It allows more water molecules to bind. Signup and view all the answers

What helps to neutralize the negative charge of the phosphate backbone in nucleic acids?

Counter ions like Mg2+ and K+, Na+ Signup and view all the answers

What type of DNA structure is a Holliday junction associated with?

Involves the exchange between adjacent helices Signup and view all the answers

Which factor is NOT associated with stabilizing nucleic acid structures?

Disruption from high salt concentrations Signup and view all the answers

Which property does Z-DNA exhibit that is different from both A-DNA and B-DNA?

Long and narrow zig-zag structure Signup and view all the answers

What is the energetic cost associated with melting DNA?

Element of entropy from conformational locking Signup and view all the answers

What role do metal ions play in nucleic acids?

They provide stability by neutralizing negative charges. Signup and view all the answers

Why is DNA in the cell predominantly in the B-DNA form?

It is the most biologically significant structure. Signup and view all the answers

What is the significance of the six single bonds between ribose molecules in the nucleic acid backbone?

They allow for rotation and contribute to the backbone's flexibility. Signup and view all the answers

Which of the following best describes the conformational variations of the deoxyribose sugar in nucleotides?

The sugar can exist as either endo or exo conformers. Signup and view all the answers

How do the nucleotide bases differ from amino acid sidechains?

Nucleotide bases are limited to purine and pyrimidine sizes. Signup and view all the answers

What type of interactions do nucleotide bases primarily utilize with ribose in nucleic acids?

Hydrophobic interactions along their flat surfaces. Signup and view all the answers

Why is it necessary to draw a Ramachandran plot for DNA in six dimensions?

To represent the six bonds with limited rotational freedom. Signup and view all the answers

is RNA ordered or disordered

ordered Signup and view all the answers

What is a characteristic of Hoogsteen base pairing?

It involves pairing of A-T and G-C. Signup and view all the answers

Which statement correctly describes the interactions in an extended triple-stranded RNA structure?

It features mixed base pairing with strand swapping. Signup and view all the answers

What type of secondary structure is formed by a 'bulge' in RNA?

A single nucleotide that is unpaired. Signup and view all the answers

How do non-canonical base pairs in RNA affect its structure?

They distort the backbone and affect spacing. Signup and view all the answers

Which of the following RNA species is NOT known for folding into complex 3D structures?

linear mRNA Signup and view all the answers

What is the significance of the major groove in Hoogsteen interactions?

It enables the insertion of additional nucleotides. Signup and view all the answers

What best describes the function of riboswitches in RNA?

To regulate gene expression by changing conformation. Signup and view all the answers

Which part of the Cro protein is primarily responsible for forming interactions with DNA?

Helix 3 Signup and view all the answers

What structural feature of the Cro protein aids in dimerization?

Beta-sheet Signup and view all the answers

How does helix 3 of the Cro protein interact with the DNA molecule?

It inserts into the major groove of DNA. Signup and view all the answers

What is the significance of the organization of the Cro dimer with respect to helix 3?

It ensures both protomers interact on the same surface. Signup and view all the answers

In the context of Cro protein's interaction with DNA, which of the following statements is true?

Only one of the half-sites is recognized by each Cro protomer. Signup and view all the answers

What role do magnesium ions (Mg2+) play in the structure of the SAM riboswitch?

They coordinate to enhance the proximity of backbone elements. Signup and view all the answers

Which interaction is indicated by the GA base pair in the SAM riboswitch?

A specific arrangement with distinct hydrogen bonds. Signup and view all the answers

What distinguishes the G.C-A base triplet interaction in the SAM riboswitch?

It represents a rearrangement of canonical base pairs. Signup and view all the answers

Match the types of amino acid residues with their characteristics in DNA binding proteins:

Arginine (Arg) = Electropositive and interacts with DNA Lysine (Lys) = Electropositive and binds to DNA Aspartic Acid (Asp) = Negatively charged and less common in DNA binding Glutamic Acid (Glu) = Negatively charged and less common in DNA binding Signup and view all the answers

Match the properties of DNA binding proteins with their effects:

Rich in Arg and Lys = Strengthens DNA interaction Poor in Asp and Glu = Reduces negative charge interactions Electropositive surface = Enhances binding affinity to DNA Electrostatic interactions = Facilitates stabilization of protein-DNA complex Signup and view all the answers

Match the terms related to DNA binding proteins with their descriptions:

Cro protein = A protein that binds to DNA to regulate gene expression Dimerization = Formation of a complex from two protein molecules Electropositivity = Characterized by a surplus of positive charge Binding specificity = Ability to selectively interact with specific DNA sequences Signup and view all the answers

Match the interactions of DNA binding proteins with their roles:

Hydrogen bonds = Stabilizes the protein-DNA interface Ionic interactions = Attracts oppositely charged groups Hydrophobic interactions = Promotes structural integrity Van der Waals forces = Facilitates close proximity interactions Signup and view all the answers

Match the aspects of protein-DNA interactions with their descriptions:

Electrostatic potential = Influences binding dynamics Surface charge distribution = Determines protein affinity for DNA Amino acid composition = Affects protein stability and function Interaction strength = Impacts gene regulatory mechanisms Signup and view all the answers

Study Notes

Nucleic Acid Structure

Nucleic acids are composed of fundamental building blocks called nucleotides
Nucleotides consist of a phosphate group, a sugar (deoxyribose in DNA, ribose in RNA), and a nitrogenous base
Purines (adenine and guanine) and pyrimidines (cytosine, thymine in DNA, uracil in RNA) are the nitrogenous bases
Nucleotides are linked together through phosphodiester bonds, forming a sugar-phosphate backbone
Bases are attached to the sugar at the 1' position.
The sugar is phosphorylated at the 5' position.

Nucleotide Organization

In DNA, the sugar is 2' deoxyribose; in RNA, the sugar is ribose
The bases are attached at the 1' position of the sugar.
The phosphate group is attached at the 5' position of the sugar.

Purines vs. Pyrimidines

Purines and pyrimidines are heterocyclic aromatic compounds.
Purines have two rings, while pyrimidines have one ring.
Biological bases are derivatives of these structures with additional keto, amine, and methyl groups.

Standard Bases

Adenine (A) and guanine (G) are purines
Cytosine (C), thymine (T) (in DNA), and uracil (U) (in RNA) are pyrimidines
Bases can be methylated on heteroatoms
In DNA, methylation is typically regulatory
In RNA, modifications (e.g., methylation in ribosomes) often play a structural role.

Deoxynucleotides

Deoxynucleotides are the building blocks of DNA
Nomenclature: Deoxyadenosine 5'-monophosphate (dAMP), deoxyguanosine 5'-monophosphate (dGMP), deoxythymidine 5'-monophosphate (dTMP), deoxycytidine 5'-monophosphate (dCMP)
Note a nucleoside refers only to the base.

Nucleic Acid Structural Hierarchy

Nucleic acids have primary, secondary, and tertiary structures
- Primary structure is the sequence of nucleotides
- Secondary structure is the base pairing interactions (e.g., double helix)
- Tertiary structure is the three-dimensional arrangement of secondary structures.

DNA Backbone and Interactions

DNA backbone is comprised of 5'OH of one ribose group linked to 3'OH of the next through a phosphate group
DNA backbone has no H-bonding donating groups, and no positively charged groups
Phosphate groups interact well with water
Deoxyribose is nonpolar
Unlike peptides, the backbone cannot make strong interactions with itself

Ribos-Phosphate Backbone

Six single bonds (alpha, beta, gamma, delta, epsilon, and zeta) exist between the 3' of one ribose and the next, allowing free rotation
Angles are highly limited by ribose ring constraints
Nucleic acid backbones have greater flexibility per residue compared to peptides.

Sugar Pucker

Deoxyribose has limited flexibility
Four atoms in the ring are planar while the other is either endo (toward C5') or exo (away from C5') for C-2' and C-3'

Base Interactions

Bases have H-bond donor and acceptor properties
Bases have a big flat, hydrophobic surface facing away from the minor groove.
Hetero-cyclic bases develop complex electrostatic fields
These fields dictate base stacking behavior
Base stacking is a crucial force in DNA stability.

Base Pairs

Watson-Crick pairs include A-T with two hydrogen bonds and G-C with three hydrogen bonds
C1-C1 distances are essentially identical at 10.9 Å in WC pairs
GC base pairs have stronger stacking than AT
Non-Watson-Crick base pairs can exist and are important for RNA structures
These non-canonical interactions distort the backbone.

DNA Double Helix

DNA exists in an antiparallel fashion
Bases interact through hydrogen bonds inside
Phosphate groups are on the outside to minimize electrostatic repulsion
The helix is right-handed

Major and Minor Grooves

The major groove is longer while the minor groove is shorter
The major groove has more room for interactions with proteins
The minor groove offers potential for sequence specific interactions to recognize the DNA backbone.

DNA Geometry

B-DNA: right-handed helix, base pairs perpendicular to the helix axis, ~23 Å diameter
A-DNA: right-handed helix, base pairs tilted, ~27 Å diameter
Z-DNA: left-handed helix, zig-zag backbone.

DNA Hydration

DNA forms hydrogen bonds with surrounding water molecules
Specific positions bind water molecules via standard interactions within the major groove or the minor groove/backbone
Water significantly contributes to B-DNA stability

Metal Ions in Nucleic Acid Structure

The large negative charge on the phosphate backbone of DNA needs counter ions (like Mg2+) to stabilize the structure
These metal ions facilitate the formation of the DNA double helix in solutions with predominantly water molecules

Forces Stabilizing Nucleic Acid Structure

Base Stacking (hydrophobic interactions, van der Waals and electrostatic).
Hydrogen bonds between bases
Entropic gain from releasing waters associated with single-stranded nucleotides into the bulk solution.
Metal ion binding

Energetics of DNA Duplex Formation

DNA duplex formation is favored under physiological conditions due to the substantial enthalpic gain from hydrogen bonding and van der Waals forces in base stacking.
Entropy of duplex formation (TDS) is unfavorable, reflecting the cost of restricting conformational freedom
DNA is readily melted by increasing temperature.

DNA Tetraplex

DNA tetraplex forms in guanine rich sequences.
Stable over a wide range of conditions.
Can be parallel or antiparallel.
Influences transcription, replication and recombination.

DNA Holliday Junction

Formed during DNA repair or meiotic crossover
Adjacent helices have complementary DNA strands exchanged
DNA flexibility allows for this configuration without significantly distorting the DNA duplex.

DNA Structure and Protein Binding

Most cellular DNA is in B-DNA conformation.
Complementary base pairing provides built-in ability for DNA to recognize complementary sequences.

Ribonucleotides and RNA

RNA is made from Ribonucleotides
Contains ribose sugar instead of deoxyribose in DNA
Uracil (U) replaces thymine in RNA
RNA generally forms A-form helical structure.

RNA Topology Diagrams

Depict the relative locations of secondary structural elements like bulges, internal loops, and hairpins

RNA Function

Function of many RNA molecules depends on folding into complex 3D structures
Including rRNA, tRNA, and mRNA in protein synthesis, as well as involved in splicing machinery.

Riboswitches

Occur in 5' untranslated regions of mRNAs in gram-positive bacteria
Control protein levels post-transcriptionally
Regulate transcription and translation depending on the presence of specific metabolites in the cell.

SAM Riboswitch

A highly complex three-dimensional RNA structure.
Contains regions that resembles standard A-RNA structures, and others with more random appearances
Many odd pairings (non-canonical interactions) in this structure.

Ligand Recognition by RNA

S-Adenosyl methionine (SAM) binding pocket involves 11 nt from 5 strands.
Interactions involving base stacking, van der Waals interactions, and multiple hydrogen bonds using both base edges and ribose atoms.
Clefts in the structure are used to stabilize SAM binding.

Hammerhead Ribozyme

Enzyme RNA strand binds and cleaves a substrate RNA strand.
Multiple Na+ ions involved in stabilizing structure.
Regions of the enzyme interact successively with the substrate RNA

Protein-Nucleic Acid Interactions

Essential for DNA maintenance, replication, and transcription.
Specific protein interactions with DNA are essential for biological processes
This include enzymatic reactions, regulation and transcription, replication.

DNAse I-DNA Interactions

DNase I is a relatively non-specific double-stranded nuclease from bovine pancreas
Binds in the minor groove of DNA, opening it up slightly
Interacts mainly with phosphate groups of the backbone through hydrogen bonds and electrostatic interactions.
Arginine residues are favored during the interactions.

Protein Targeting

There are 4 bases in DNA (A, G, C, and T).
A set of n nucleotides has 4n possible sequences.
In the human genome, this is a very large number.

"Unwind and Read" Strategy

Proteins may possibly unzip DNA to "read" the base pairing edges.
This strategy is energetically expensive.
However, some proteins use modified versions of this strategy exploiting chemical modifications of bases to flip a base out of the DNA

Recognising Base Edges

This strategy is plausible because each base pair has a unique pattern of potential interactions in the major groove.
The minor groove is more ambiguous (difficult to interpret).

Helix-Turn-Helix Transcription Factors

Recognition helices in these types of proteins often insert into major groove of DNA.

CRO Protein and DNA Binding

Recognizes a 17 residue pseudo-palindromic sequence.
Forms a dimer.
Helix 3 for both protomers sticks out on the same surface interacting with DNA half site.

Cro Interactions

helix 3 inserted into DNA major groove
Hydrogen bonds mediated with individual bases
Non-specific interactions with backbone, dominated by basic residues (Lys/Arg) and electrostatic interactions.
DNA is distorted upon binding to form a specific interaction site between the protein and the DNA

HTH Motif in DNA Binding Proteins

Recognition helix in DNA-binding proteins inserts into major groove and interacts specifically with bases
Helix near the N-terminus position is responsible for recognition helix position and orientation

Other Non-HTH Proteins

Some proteins also use a-helices to recognize DNA (e.g. coiled-coil, zinc fingers or leucine zipper)

TATA Binding Protein (TBP)

Ubiquitous eukaryotic transcription factor.
Directly binds to DNA TATA box, assisting in recruitment of RNA polymerase.

TBP Structure

Monomeric protein with structurally similar halves.
Shaped like a saddle.

TBP DNA-Binding

Binds in the shallow minor groove.
Contacting 8 consecutive base pairs.
Extensive interactions with the DNA backbone using basic amino acid residues.

TBP DNA Contacting Bases, Additional interactions

TBP forms specific hydrogen bonds with AT edges using basic residues.
Extensive interactions with the DNA backbone due to interactions with the PO4 groups
Surface contacting DNA bases is primarily composed of aliphatic and aromatic residues, contributing to the overall hydrophobic nature of the protein-DNA interface

Modular DNA Recognition by TAL Effector Proteins

TAL proteins are built from sequential repeats.
Each repeat comprises two helices.
This modular design makes them highly customizable for protein engineering.

K16 and Q17 of Each Repeats Contact PO4

K16 and Q17 amino acids contact the phosphate group of the DNA strand.
These contacts provide high binding energy without sequence specificity

Positions 12 and 13 Specifying the Base

Position 12 (Asn or His) and 13 (Gly, Asp, Asn, or Ser) residues together determine nucleotide specificity.

RNA Topology diagrams(additional details)

Depicts secondary structural elements like bulges, internal loops, and hairpins.

Disorder-Order Transition in NA Binding

DNA binding proteins often have disorder.
Binding to DNA partner forces the protein into ordered conformation.
Large number of basic amino acids and non-polar base stacking observed.

General Conclusions

DNA binding proteins primarily interact with phosphate backbone to derive their binding energy.
Almost all sequence-specific DNA-binding proteins distort DNA.
Specificity is heavily influenced by the DNA's ease of twisting/distortion into the specific conformation for binding
Hydrogen bonds with exposed bases are essential for sequence recognition.

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Description

This quiz explores essential concepts related to DNA binding proteins, including their structural features, interactions with DNA, and the significance of hydrophobic interactions and ordering transitions. Test your knowledge on topics like the major groove's role in recognition, the phosphate backbone, and the mechanics of DNA distortion.