BG1141 Cell & Molecular Biology for Bioengineers - Protein PDF

BG1141: Cell & Molecular Biology for Bioengineers Protein The building block Structure and folding Proteins in Everyday Life http://www.answerfitness.com/ Proteins in Our Body...

BG1141: Cell & Molecular Biology for Bioengineers Protein The building block Structure and folding Proteins in Everyday Life http://www.answerfitness.com/ Proteins in Our Body Keratin (hair, nails, skin) Collagen (skin) Hemoglobin (blood) https://s3.amazonaws.com/ http://www.healthyprotocols.net http://www.eternogen.com/media/collagen-diagram.jpg General Protein Functions  Correct structure and conformation are required for proteins to function correctly The correct structure and conformation depends on protein folding Protein } Consists of one or more chains of amino acids } Translated from RNA 5’ to 3’ that corresponds to N- and C-termini } Performs specific function } Some undergoes post-translational modification (e.g. addition of sugar) } Has 4 hierarchical structures } Primary (the amino acid sequence) } Secondary (resulting from the interactions between amino acids) } Tertiary (resulting from the interactions between 2o structures) } Quaternary (resulting from the interactions between 3o structures) 6 Central dogma of molecular biology DNA RNA protein 7 The flow of genetic information in cells is from DNA to RNA to protein Genetic information directs the synthesis of protein From DNA to Protein Folded protein Image sources: www.genome.gov & nobelprize.org DNA and Protein Sequences } DNA = nucleotide or nucleic acid sequence LOCUS E2WT_CODA 783 bp DEFINITION E2WT_CODA 780 bp 5-->3 + taa (stop) ORIGIN 1 ATGCTGTCTG TTCCTGGTCC CGCTGCTGCA GAGGAAAAGG CTGCTCCAGC GGCTGCGAAA 61 CCGGCTACTA CTGAAGGTGA ATTCCCTGAA ACCCGTGAAA AAATGTCTGG TATCCGTCGT 121 GCAATCGCGA AAGCCATGGT TCACTCTAAA CACACCGCGC CACACGTTAC CCTGATGGAT 181 GAAGCAGACG TTACCAAACT GGTTGCGCAC CGTAAAAAAT TCAAGGCGAT TGCGGCGGAA 241 AAAGGTATCA AACTGACCTT CCTGCCGTAC GTTGTTAAAG CTCTGGTTTC GGCTCTGCGT 301 GAATACCCGG TTCTGAACAC CTCTATTGAC GACGAGACCG AAGAAATCAT CCAGAAACAC 361 TACTACAACA TCGGTATCGC TGCGGACACT GATCGTGGTC TGCTGGTTCC TGTGATTAAA 421 CACGCGGACC GTAAACCGAT CTTCGCGCTC GCTCAGGAAA TCAACGAACT GGCTGAGAAA 481 GCTCGTGACG GTAAACTGAC TCCTGGTGAA ATGAAAGGCG CGTCTTGCAC TATTACCAAC 541 ATCGGCTCTG CAGGTGGTCA GTGGTTCACC CCAGTTATCA ACCACCCGGA AGTTGCGATC 601 CTGGGTATTG GTCGTATAGC CGAAAAGCCG ATCGTTCGTG ACGGTGAAAT CGTTGCTGCT 661 CCGATGCTGG CCCTGTCTCT GTCTTTCGAT CATCGTATGA TTGATGGCGC GACCGCACAG 721 AAAGCCCTGA ACCACATCAA ACGTCTGCTG TCCGACCCGG AACTGCTGCT GATGGAAGCT 781 taa } Protein = amino acid sequence 1 11 21 31 41 51 | | | | | | 1 MLSVPGPAAA EEKAAPAAAK PATTEGEFPE TREKMSGIRR AIAKAMVHSK HTAPHVTLMD 60 61 EADVTKLVAH RKKFKAIAAE KGIKLTFLPY VVKALVSALR EYPVLNTSID DETEEIIQKH 120 121 YYNIGIAADT DRGLLVPVIK HADRKPIFAL AQEINELAEK ARDGKLTPGE MKGASCTITN 180 181 IGSAGGQWFT PVINHPEVAI LGIGRIAEKP IVRDGEIVAA PMLALSLSFD HRMIDGATAQ 240 241 KALNHIKRLL SDPELLLMEA Number of amino acids: 260 9 Molecular weight: 28116.6 Theoretical pI: 7.14 From DNA to RNA = transcription Genes can be expressed with different efficiencies. Many identical RNA copies can be made from the same gene, enabling 10 rapid synthesis of protein. From RNA to protein = translation } Translation } Conversion of the information in RNA into protein } Genetic code } Set of rules specifying the correspondence between nucleotide triplets (codons) in DNA or RNA and amino acid in proteins } Codon } A group of three consecutive nucleotides in RNA } Each codon specifies one amino acid } Reading frames } The set of successive triplets in which a string of nucleotides is translated into protein. } An mRNA molecule is read in one of three possible reading frames, depending on the starting point. 11 Proteins are translated by polyribosomes (polysomes) – large cytoplasmic assemblies made of several ribosomes spaced as closely as 80 nucleotides apart along a single mRNA molecule 12 The nucleotide sequence of an mRNA is translated into the amino acid sequence of a protein via the genetic code wobble 13 An RNA molecule can be translated in three possible reading frames 14 Amino acid is the building block of protein } Each translated from a codon (3 nucleic acid) } Approximate molecular mass of each amino acid: } 110 Da (1 Da ~ 1 g/mol) } Each amino acid has } amine (-NH2) group } carboxy (-COOH) group } residue (R; also called side) group } The residue group determines the characteristic of each amino acid } Alpha carbon a-carbon } Where the functional group is attached to 15 Amino acid is the building block of protein } All amino acids derived from protein will have the L- stereochemical configuration (L-amino acid) } The other is D-amino acid, usually component of short bacterial peptide } 20 standard amino acids } Amino acid derivative, e.g.: GABA, etc. } Voet p. 86 16  17 Source: Bachem webiste https://www.bachem.com/fileadmin/user_upload/pdf/Catalogs_Broschures/PCAA.pdf One-letter code Three-letter code Full name To remember A Ala Alanine C Cys Cysteine D Asp Aspartic acid AsparDic E Glu Glutamic acid GluEtamic / GlutEmic F Phe Phenylalanine Fenylalanine G Gly Glycine H His Histidine I Ile Isoleucine K Lys Lysine LyKesine L Leu Leucine M Met Methionine N Asn Asparagine AsparagiNe P Pro Proline Q Gln Glutamine Qlutamine R Arg Arginine Rginine S Ser Serine T Thr Threonine V Val Valine W Trp Tryptophan Twiptophan / 2 rings, 2 Vs 18 Y Tyr Tyrosine TYrosine Some unique features } Ala: the smallest side chain } Gly: the smallest AA } His: contains imidazole group } Pro: found in turns } Cys: contains sulfhydryl (-SH) group } Side chains that are: Aromatic Trp, Phe, Tyr Amino-containing His, Arg, Lys Carboxyl-containing Asp, Glu Hydroxyl-containing Ser, Tyr, Thr Thiol-containing Cys, Met 19  Amino acids are covalently linked through peptide bond The C-terminus from one connects to the N-terminus of the next Peptide bond Condensation reaction Resonance form of peptide groups Page 127 http://www.youtube.com/watch?v=Vvm-wK2vSO8 Source: wikipedia Figure 6-2  Polypeptide is formed from a chain of amino acids } Monomer = amino acid } Polymer = peptide à polypeptide } Peptide is short sequence ( pI à net charge is negative } As pH increases, acidic and basic groups are deprotonated } Carboxyl à carboxylate anions (R-COOH à R-COO-) } Ammonium à amino (R-NH3+ à R-NH2) } pH < pI à net charge is positive } Most proteins have pI between 5 and 8.5 } pH, pK, and charge of amino acid residue are related by Henderson- Hasselbalch equation [𝑐𝑜𝑛𝑗𝑢𝑔𝑎𝑡𝑒 𝑏𝑎𝑠𝑒] 𝑝𝐻 = 𝑝𝐾 + 𝑙𝑜𝑔 [𝑐𝑜𝑛𝑗𝑢𝑔𝑎𝑡𝑒 𝑎𝑐𝑖𝑑] 47 Antibody: the IgG } Consists of 4 hetero subunits } Light chain = 2 } Heavy chain = 2 } Fab or Fv = antigen-binding fragment or variable fragment } Fc = constant fragment } Antigen binding site has Fab and Fc 48 Figure 7-39 Figure 7-40 Allostery } Allosteric Effects (p.192): } binding of a ligand at one site affects the binding of another ligand at another site, generally require interactions among subunits of oligomeric proteins. } Involves slight change of protein conformation } Example: binding of oxygen to haemoglobin } Most enzymes are not allosteric 51 Sequential model proposed by Daniel Koshland Table 5-1  Protein Data Bank (PDB): http://www.pdb.org Protein parameters on ExPASy: http://web.expasy.org/protparam/ Table 5-4 Learning Points } Protein is made of amino acids } The interaction between amino acids results in 1°, 2°, 3°, 4° structures } Folding is determined by the sequence until it reaches thermodynamic equilibrium } The conformation is stabilized by different forces } The structure can be determined using X-ray crystallography and NMR } Parameters that describe proteins } Allostery effect is the change of protein conformation due to binding of a certain molecule 55