Experimental Methods Class Notes June 6, 2001

Class Notes: June 6

Mutations and SNPs: we use SNPs for Mappin !enot"pin : to identif" indi#idual enomes P$%0s are a famil" of en&"mes in the li#er that modif" dru s and determine whether a dru will wor' in the human meta(olism or not) *ou +an impro#e the odds of dru su++ess (" +omparin results to people with the ,+orre+t- SNP at the p$%0 of +hoi+e) S+reenin is +arried out (" +on#entional methods S+annin lar e populations is done (" mi+roarra"s, PC., or se/uen+in )

106 possible compounds

1000 possible compounds Today the bottleneck is here, and its Caused by metabolism

100 possible compounds

Microarray screening of SNPs

!enomi+ 1N2

PC.

PC. fra ment: re ion of interest

S"stemati+all" +han e ea+h oli o (" one (ase pair)

Why cant you unboil an egg? Can proteins repli+ate0 No, (ut the" +an +ause other proteins to unfold)

Experimental Methods Class Notes June 6, 2001 Stanle" Pruissner is a s+ientist who dis+o#ered prions, the unfolded proteins that sometimes +ause disease li'e Mad Cow) 1a#id Eisen(ur is a 3C42 professor who wants to ma'e a data(ase of fun+tionall"5 related proteins) Protein Folding Su+l"+in is a protease +ommonl" found in deter ents and +onta+t lens solutions) Chaperones e) ) 6eat Sho+' Proteins 76SP8 that a++elerate protein foldin ) Protein synthesis Good test question: 6appens in the ri(osome Can proteins s"nthesi&e from (oth 4in'ed (" peptide (ond 7C5N8 N5terminal to C5 2lwa"s unidire+tional: from N5terminal to C5terminal) terminal0 9he primar" ri(osome produ+t is a pol"peptide +hain) o 2lwa"s 1 ene 1 m.N2 s"nthesis of pol"peptide +hains o 20 natural amino a+ids 6"drophili+ 6"dropho(i+ 2+idi+ :asi+ Sulpher5+ontainin o Non5natural amino a+ids Can (e used as pro(es Perhaps in spe+ial dru s) Protein Structure Primar": the amino a+id se/uen+e) Se+ondar": the ;1 stru+ture, <0= of whi+h is made up of >5heli+es, ?5sheets, and re#erse turns) 9ertiar": the ;1 +onfi uration and how it fits with other proteins) @uaternar": intera+tion (etween pol"peptide +hain su(unitsA for example, homodimers 7sin le su(units8 and heterodimers 7multi5su(units8) 1omains: the tertiar" or se+ondar" stru+ture Compa+t lo(ular unit whi+h +ontains the fun+tional part of a protein) Ma" point toward eneti+ and +ellular heredit" of proteins) Motif: se/uen+es not asso+iated with stru+ture Pol"peptide se/uen+es that are asso+iated with fun+tion) Note: domains and motifs are shared a+ross a famil" of proteins)

Experimental Methods Class Notes June 6, 2001 *ou +an predi+t motifs from se/uen+es, (ut it is hard to predi+t domains) Post translational !odifications Proteol"ti+ pro+essin 7e) ) (lood +lottin 8 o Bor se+retionleader peptides o Cna+ti#e to a+ti#e en&"me: &"mo ensinsulin Proteol"ti+all" pro+essed, these en&"mes ha#e different +ellDor an sour+es) o Eiral assem(l": #iral m.N2 to protein is pro+essed (" #iral proteases) o Pol"peptide spli+in : +an+a#alin 2, final pol"peptide is a spli+ed #ariant of the ori inal) o 2lteration of +hain termini N5terminal met is atta+hed with a form"l roup) 2+et"lation of N5terminus !l"+os"l5phosphate mem(rane an+hors atta+h to +ell mem(ranes of solu(le proteins) 9he followin t"pes of +han es +an (e predi+ted: !l"+os"lation o N5lin'ed: mostl" in E).) and on 2sn o F5lin'ed: SerD9hr in ol i (odies 4ipid atta+hment: to tether solu(le proteins to mem(ranes o E) ) m"ristol, palmito"l, et+) Phosphor"lation o Primaril" at F6 roups of Ser, 9hr, or 9"r 1isulphide (ond formation: (onds (etween two +"stines Why deter!ine protein structure? Stru+ture determines fun+tion 1enotes relationships (etter than se/uen+e Small mole+ules often alter protein fun+tion (" alterin lo+al stru+tures