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Recommend the majority of chapter 3Proteins and Amino Acids1

2Section 2: amino acids and proteins1) Amino acidsZwitterionsAmphoteric: acid/basebehaviour of amino acidsNon-ionizable and ionizable R-groupsTitration curves of amino acidsIsoeletric point, pIAmino acid classificationAmino acid stereoisomerism

2) Amino acid ionization

3) The peptide bond

Molecular weight of AA and peptides

Peptides, Polypeptides and proteins4) Prosthetic groups

Proteins are polymers of amino acids5) Methods to study proteinsFractionationColumn chromatographyBeer-Lambert LawElectrophoresisProtein sequencing3

LysozymeProteins are the workhorses of cells: complex, convoluted, beautiful shapes that do something. Catalysis:enolase (in the glycolytic pathway)DNA polymerase (in DNA replication) Transport:hemoglobin (transports O2 in the blood)lactose permease (transports lactose across the cell membrane) Structure:collagen (connective tissue)keratin (hair, nails, feathers, horns) Motion:myosin (muscle tissue)actin (muscle tissue, cell motility)

Human genome encodes > 20,000 proteins4

Examples of protein function and structure that you can seeFireflies express the protein luciferin

www.rcsb.orgRed blood cells express the protein hemoglobin

Rhinoceros horn made of the protein keratinwww.rcsb.orghttp://www.leeds.ac.uk/heritage5

Proteins are polymers of amino acidsProteins are heteropolymers of 20 common amino acidsAmino acids have properties that are well suited to carry out a variety of biological functions:Capacity to polymerizeUseful acid-base propertiesVaried physical propertiesVaried chemical functionality6All amino acids have a common, general structure7

7Classification of the 20 common amino acidsCommon amino acids can be placed in five basic groups depending on their R substituents: Nonpolar, aliphatic (7) Aromatic (3) Polar, uncharged (5) Positively charged (3) Negatively charged (2)

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Glycine is smallProline is cyclicalThese amino acid side chains absorb UV light at 270-280 nm10

Form hydrogen bondsCan form disulphide bonds with other cysteines1112

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Amino acid short handAmino acids are represented by two short-hand codes:

Name three-letter code one-letter code 14GlycineGlyGAlanineAlaAProlineProPPhenylalaninePheFTyrosineTyrYSerineSerSCysteineCysCAspartateAspD

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16Amino acid nomenclature and classification: What do I need to know for the exam?For all 20 amino acids:

1. Given an amino acid (name OR structure), determine if it is polar uncharged, charged, non-polar aliphatic or aromatic2. The corresponding three-letter and one-letter abbreviations.3. Draw the structure of the amino acid4. Name the amino acid given the structure16Most -Amino Acids are ChiralNon-superimposable mirror image stereoisomer = enantionmerIs there an exemption?Yes, GlycineThe -carbon has always four substituents and is tetrahedral17

Amino acid L- and D- enantiomersAll a-amino acids (except glycine) have an L- and D- enantiomer18

Amino acid L- and D- enantiomers (2)L and D-enantiomers are chemically indistinct

The difference is rotates polarized light differently

In vitro synthesis of amino acids: 1:1 ratio of L to D (racemic mix)

In cells: amino acids are predominantly of the L-enantiomer

Why? 19Ionization of Amino Acids20

The carboxyl and amino groups can act as an acid or base.

Molecules that can be a base or an acid simultaneously= amphoteric

At physiological pH (7.2), the carboxyl group donates H+ while the amino group accepts a H+

At pH 7.2 amino acids are dipolar ions = zwitterions

Despite charges, at pH 7.2, these amino acids are neutral (but the R group will matter)

Ionization of Amino Acids21

Amino acids as zwitterions and amphoteric moleculesIf high pH, H+ is released

Becomes negatively chargedIf low pH, H+ is acquired

Becomes positively chargedAt pH 7.222Ionization of Amino AcidsAt acidic pH, the carboxyl group is protonated and the amino acid is in the cationic form

At neutral pH, the carboxyl group is deprotonated but the amino group is protonated. The net charge is zero; such ions are called Zwitterions At alkaline pH, the amino group is neutral NH2 and the amino acid is in the anionic form.23Chemical Environment Affects pKa Values24

What about amino acids with ionizable R-groups?25For Glutamate, which charge is predominant at pH 7?26

26For histidine, which charge is predominant at pH 7?

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How to predict pI of an AA with an ionizable chain? 28How to predict pI of an AA with an ionizable chain?

29Joining amino acids to make polypeptides

THE PEPTIDE BOND30

Formation of the peptide bond PEPTIDE BONDHydrolysis rxnCondensation rxnAmino acids in polymer are called residues3131

Peptides and PolypeptidesUse 3-letter or 1-letter code to represent sequenceSGYALAmino Acids, peptides, methods3232Peptides: A Variety of Functions Hormones and pheromones: insulin (think sugar) oxytocin (think childbirth) sex-peptide (think fruit fly mating) Neuropeptides substance P (pain mediator) Antibiotics: polymyxin B (for Gram - bacteria) bacitracin (for Gram + bacteria) Protection, e.g. toxins amanitin (mushrooms) conotoxin (cone snails) chlorotoxin (scorpions)

3334Example of a peptide: Aspartame

Aspartame is a dipeptide(L-Aspartyl-L-phenylalanine methyl ester)

N+H3CCONCCOHHHCCH2OO-CH2OHCH2O35Example of a peptide 2: insulin

Insulin is first made in cells in the pancreas as part of a longer polypeptide called pre-proinsulin, 110 amino acids longThe finished polypeptide insulin is 51 amino acids longwww.rcsb.org36Example of a peptide 3: amanitin

en.wikipedia.orgwww.mykoweb.com

www.rscb.orgamanitinRNA polymerase IIAmanita muscariaAverage Molecular Weight of PeptidesChapter 3.2 Peptides & ProteinsAverage MW of AA is 138 Da

In biology use: 128 Da (smaller AA predominate in cells)

In proteins, average MW for residues: 128 - 18 (MW H2O) = 110 Da

Therefore the #residues in a protein = MW protein / 110

Or the MW of a protein = number of residues X 110 DaAmino Acids, peptides, methods3738Question: For the peptide shown below, provide the following:

Molecular weight (based on the average molecular weight of an amino acid in a peptide) The charge on the molecule at pH 7.0The isoelectric point

N+H3CCONCCOCCONCCONCCONO-pKa = 2.34pKa = 9.60HHHHHHHH(CH2)4N+H3pKa = 10.5CH3CH2OHCH2OHHCCH2OO-pKa = 3.739The amino acid sequence of a polypeptide is its primary structure

39The amino acid sequence of a polypeptide is its primary structure (2)Primary structure of Clathrin heavy chain 1675 amino acids long40maqilpirfq ehlqlqnlgi npanigfstl tmesdkfici rekvgeqaqv viidmndpsn pirrpisads aimnpaskvi alkagktlqi fniemkskmk ahtmtddvtf wkwislntva lvtdnavyhw smegesqpvk mfdrhsslag cqiinyrtda kqkwllltgi saqqnrvvga mqlysvdrkv sqpieghaas faqfkmegna eestlfcfav rgqaggklhi ievgtpptgn qpfpkkavdv ffppeaqndf pvamqisekh dvvflitkyg yihlydletg tciymnrisg etifvtaphe atagiigvnr kgqvlsvcve eeniipyitn vlqnpdlalr mavrnnlaga eelfarkfna lfaqgnysea akvaanapkg ilrtpdtirr fqsvpaqpgq tspllqyfgi lldqgqlnky eslelcrpvl qqgrkqllek wlkedklecs eelgdlvksv dptlalsvyl ranvpnkviq cfaetgqvqk ivlyakkvgy tpdwifllrn vmrispdqgq qfaqmlvqde epladitqiv dvfmeynliq qctaflldal knnrpsegpl qtrllemnlm hapqvadail gnqmfthydr ahiaqlceka gllqralehf tdlydikrav vhthllnpew lvnyfgslsv edsleclram lsanirqnlq icvqvaskyh eqlstqslie lfesfksfeg lfyflgsivn fsqdpdvhfk yiqaacktgq ikevericre sncydpervk nflkeakltd qlpliivcdr fdfvhdlvly lyrnnlqkyi eiyvqkvnps rlpvviggll dvdcsedvik nlilvvrgqf stdelvaeve krnrlklllp wlearihegc eepathnala kiyidsnnnp erflrenpyy dsrvvgkyce krdphlacva yergqcdlel invcnenslf kslsrylvrr kdpelwgsvl lesnpyrrpl idqvvqtals etqdpeevsv tvkafmtadl pneliellek ivldnsvfse hrnlqnllil taikadrtrv meyinrldny dapdianiai snelfeeafa ifrkfdvnts avqvliehig nldrayefae rcnepavwsq lakaqlqkgm vkeaidsyik addpssymev vqaantsgnw eelvkylqma rkkaresyve telifalakt nrlaeleefi ngpnnahiqq vgdrcydekm ydaakllynn vsnfgrlast lvhlgeyqaa vdgarkanst rtwkevcfac vdgkefrlaq mcglhivvha deleelinyy qdrgyfeeli tmleaalgle rahmgmftel ailyskfkpq kmrehlelfw srvnipkvlr aaeqahlwae lvflydkyee ydnaiitmmn hptdawkegq fkdiitkvan velyyraiqf ylefkpllln dllmvlsprl dhtravnyfs kvkqlplvkp ylrsvqnhnn ksvneslnnl fiteedyqal rtsidaydnf dnislaqrle kheliefrri aaylfkgnnr wkqsvelckk dslykdamqy aseskdtela eellqwflqe ekrecfgacl ftcydllrpd vvletawrhn imdfampyfi qvmkeyltkv dkldaseslr keeeqatetq pivygqpqlm ltagpsvavp pqapfgygyt appygqpqpg fgysm

Roth TF and Porter KR, 1964Endocytosis in the yolk sack

Clathrin heavy chain (3 polypeptides)

Cysteine residues can form disulfide bonds41