BIS 102-Hilt-Final

BIS 102-Hilt-Final - Biological Sciences 102 Name Winter,...

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Unformatted text preview: Biological Sciences 102 Name Winter, 2004 ' " r " .1. Hilt Isa pl 7 C, v t Final Exam Score (200): W ‘/ pH = pK + log {[base]/[acid]} Kb = XZ/y-X AG 2 AH — TAS v5 = {Vmax [S] } / {Km + [S] a ‘ V l. You are going to measure enzyme activity of enzyme “Z”. Enzyme “Z” catalyzes the reaction: . ' s+ <—> P + nzyme “Z” has a Km = 2.5 x 10'6 M for 3*. You mix the following solu ions together: 2.60 ml 10 mM phosphate buffer, pH 7.6 0.30 ml 400 mM s+ 0.10 ml enzyme . . om manor. lbw—v \jts (“5 The results of this enzyme assay are plotted at the right: \{ [ma-«m 30 Max "Vo (5 pts.) a) How many International Units (I.U.’s) ‘of enzyme “Z” were present in the assay? Show all calculations. - (a N: am miafmta 35%;?“ Gag“) g igxggww I 1'1 i (10 pts.) b) Calculate what Vo Would be if the initial substrate concentration 0 1o 20 3° 4° so had been 1.5 x 10’6 M. Show all calculations \[WX v.” V0 1 5 Time (seconds) a (a {it WIO‘VMD figxufla a V0: rm- w Mt UM ‘ @ tram“! +v Lbyw‘tvx 44 No"! ,l 8 fl 8 “<3 nmol P produced in assay if? {(20 pts.) 0) How long (in minutes) would the reaction in part “a” above have to run before the pH was 7.50? M The pKa’s of phosphate are 2.1, 7 .2, and 12.3. Show all calculations. M m G, p.555“ fig; 3:"? ’lDi’le Me? 3.11 9m] ‘3?» Imam? Wang “fills-‘3 :52" T” Fog} Oilwélf&g 'é'iJz’ef first}? it I - A‘N :d'll bfi‘cfi \l/ ' a: . 16! WW (- 1H3 ‘3 *1 mm WW1 mm Kama (.2533 == M MA We; Meaty w if X ‘ gwskuck. 2of4 i’i' 102 _ Wm offer the greatest mobili is molecule,__it1 a membrane bilayer in a giant. Explain, on a molecular basis, why you chose the fatty acids that you did (i.e. why are these particular fatty acids so mobile?) 0(_. . . . SWCWC‘ a “if- orig/WV? antimalde bit, Fr 18 Ltmminewl/Mtl +Nrefis’c n ,1, (S -‘ ~. ' . /’\ c—o—fH g ,1“! W3 Tin-Le: :Flvudltj cute: '1:- Q) leo,_9 ,o_%__CtH amen/cacti 1W5 W ('2’ cm- Exiizeataaa‘efsswm W M W News were” WWW) ‘ dim Jo gm ems LO/wwc‘h‘rflvk “R (566 abflf) .C . I I c (_23 WEEK ‘iwu (fibre atSD Jena. er'm .F.“ ohkhqs in rectors? mmbmhu, 3. (16 pts.) Appropriate chemical standards are an absolute necessity in evaluating experimental results. Fill in the empty boxes in the following table: A/ U» i iochemical Techni ue, Procedure, or Machine Standards _ 1. Amino acid anal sis 1. .20 free amino acids Protein seuenator -‘ v-r - ‘ 2. TC-a ‘ e a r . .A M lg 3. fa acid meth l esters 4. Proteins of known MW treated with sodium (’59 5‘) dodec l sulfate, merca toethanol, and heat 5. Saner metho ' . . . . .‘ . . . _ ‘xt'_.l. 'W,CJ-- 4. (21 pts.) Fill in the missrng values in the followmg enzyme punfication table. .{ 1 . ,. .1 rt / e» Volume [Protein] Total Activity Specific Fold Fraction m ml) LUJml I.U. Yield a Purification WEI-m- 2. resuspended 200 20 12 moo 120 U 3 40% (NI-14)ZSO.. I ‘ llet 29:7 10%)?) l . 200: f, ego—g J ’ 1—H K’ . ou want to purify enzyme Z. Enzyme Z has never been purified before. (10 pts.) a) What assay(s) will you need in order to follow the progress of enzyme Z’s purification at each step? Explain the chemical basis of your assay(s) and why you need them. ark » . a a . . I. h a - a sat +v War we $o€£~fi " r: awaax , (a cwWall m w: an _ \ t e" 9% M; 5. Wu km.“ W Lavage) M p a (esterztdrwfi +9 feel-"WW Mars Wt Wm :5va (win ge+ug>~§¢¢m_ MM ~5v‘ke (21%“. gammzmbw (10 pts.) b) Let’s say that you have the following techniques at your disposal. Rank them from “hig ” to ’ “low”, in terms of which technique will most likely give the greatest fold purification of your enzyme Z. Technigre Sizing column {NI—102804 cuts ' IBF gel Cation exchan e column 4" Affinity olumn _ 4 M8? Me amen/m s (10 pts.) 0) How will you know when you are done purifying enzyme Z? If there is more than one criterion, then list them all. - Rank (1 = best technigflie, in terms of fold purification; 5 = worst technique) { 3s. p wit Wflh W, specific fifirgfifi immstxfi W {nigh 53m? vying mefggflfim * U ./ n , . refit/Y W mam “were. \5 emu rant brawl (me \V l 8‘ \J/ (20 pts.) Approximately how many hydrogen bonds (fill in each blank with a number) are present in each of these 2° protein structures? (Cf—«i “ ” 3- a ‘. +3a) an oc-helix that has five complete turns ‘ NM .l/L ( V a a W hm) ,, 3g}? Lfb) a reverse turn ’ 2— I + éc) two strands of antiparallel B-pleated’sgeet; eacE in? nd 5 amino acid residues 3 7. (15 pts.) AS is an important player in biochemistry. Explain how AS is involved in each of the following ’ mical concepts: 1 l0 . o a) protein 3 structure: + A$ 9-? 1WD 56$ We Men the Wain «tame {dads-f firm aw, twmgflwfic iféfiffl W‘Wmfi 5 Wéiorté the “meaan r :3 at?" traumas»: castrate? a: l r 53 «"13 " _. 4‘ 25b) em%agc:tal£i§2 . (W M19» as OHMGW damage W o m magma damages 53$qu by Maya l5? s untrue 5% Owl bihcli‘hg “k2 data; Sub smut {gums ‘ f c) f i 1” ‘ u , (identify a topic discussed in our course, that involves ' not related t the other t 0 already mentioned). You do not have to discuss this one, just identify it. 2 \y s 8. Compound “I” trims out to be inhibitor of enzyme Z. (7 pts.) a) Put arrows (2) into the appropriate box(es) in the diagram below: E+S ES —> E+P @‘i i |:l EI + s D ESI _ a (8 pts.) b) Explain what effect, if any, this uncompetitive inhibitofillas on Km and Vm and why: Effect on Km (and why): 5:“ kc? ‘97“ pm *4 M 3,6 m rm N5 alrech 1' g EHF’E. S JIYQCfis‘u‘ +cwttrd~+ "\ . WWW? - {naming 1 MW mum lessoF 0n O'FE‘WS 6 _ Effect on Vm,‘ (and why). bk ,1, any, ewfi a“; u 1 r ‘1“ .55» comm: has Mei ; “*3 {4 VWX b’& a: at?!" W {S 3mg; mg}qu We (n+3 oppmg... Maison less 41mm Mal—1+- wma be 14 4+“ waS m, :1; 9. (6 pts.) Draw the complete structure of the largest amino acid, in the form that would predominate at pH 7: *_ 1 _ __ ; Have we . é) . 0H1. H J')‘ H \ Hgfi,fi,—Coo: “EN-§L.-coo. C9 w: r ., H-—"’<:E:o/ :3; +NH3 aft)? .w'n'um Km Hr<-..____'__Hfi7' (e) 1 1. (15 pts.) What is the final pH of a solution obtained by mixing 200 ml of 0.400 M NH3 with 300 ml of 0.100 M HCl? (K, = 1.8 x 10's) Show all calculations. (Writ (“W Nels) Qcomr dMflmia dMfian’smorHu AH 0.61M lqu‘F-K «finial-pr ‘I X Biological Sciences 102 Name VFall, 2001 Last, First K. Hi]: Final Exam Instructions: You may use the back of these pages if you clearly indicate that you are doing so. A straightedge has been provided for your use in graphing. — — “— — n- — - Total: 200 — *I-I —— — _ _ ma- _-EIE- _ _ .m- Olioetide N-terminal amino :4 cu Olioetide C—terminal carbo 13 n “a pH = -log [H*] pH = -log am [(3 = x2!(y — x) pH = (pKai + pKa2)/2 pH = pKa + log {[base]/[acid]} (Ka) (Kb) = 1.00 x 10‘” Kb = x2! (y-x) [I G = DH - TDS F = (q1 qz)/(e r2) a 7 0 Va: {Van-x [3]} 2' {Km + [3]} Biological Sciences 102 1. Let's take a look at the amino acid glycine. ( 3 pts.) a) Draw the structure of glycine in the form that would predominate at pH 7: O l H (6 pts.) b) Over what pH range would glycine bind to an anion exchange resin? Show any calculations that you may do to support your answer. (12 pts.) c) Glycine residues, along with proline residues, are often found as components of this type of 2° protein structure. Draw a portion of this 2" structure, using atoms, including the pro and gly residues in their - proper location. {Amuse *W W d) Glycine is usually the trailing ion in native and SDS-PAGE ads Glycine is deliberately put into the upper reservoir buffer in the electrophoresis. Glycine takes part in the flaking process that reduces large sample volumes to very thin lines in the stacking gel. The pH of the uppa reservoir bufi‘er is 8.3, while the pH of the stacking gel is 6.8 and the pH of the running gel is 8.8. The positive ductrode is at the bottom of the gel. ( 7 pts.) #1) Do the glycine molecules speed up, slow down, or move at the same speed as they migrate into the stacking gel from the upper reservoir? Explain S’Va‘fl We SaWQ \Oi L don‘ 't MUC QVMXVV V. ‘6') Q ( 2 pts.) #2) Do the glycine molecules speed up, slow down, or move at the same speed as thq migrate into the running gel from the stacking gel? Explain Biological Sciences 102 Name 2. (10 pts.) A protein was completer digested with trypsin and an internal pentapeptide (i.e. not derived from the N- or C-terminals) was purified. Determine the pentapeptide's sequence from the following information: a) treatment of the intact pentapeptide with chymotrypsin yields a dipeptide containing phenylalanine and a tn'peptide; b) treatment of the intact pentapeptide with the sequence of Edman chemistiy steps yields PTH-leu; and 0) treatment of the intact pentapeptide with cyanogen bromide yields free methionine, free arginine, and a tripeptide. \'( The sequence of the pentapeptide is: l6; 5 Q“ 6 mo): vhf-33R cw gt (give the three letter code of each amino acid resi ue) ‘ 3. Weak bonds are very important in biochemistry. ( 5 pts.) 3) Which weak bond is predominant in detennining fluidity between neighboring fatty acid chains in a membrane? a \lfi v“ okw WM l ( 5 pts.) b) Which weak bond is predominant in forming 3" protein structure? p.45 (opt/10b" C‘ ( 5 pts.) c) Which weak bond is the major form of communication between subunits in hanoglobin? it l' ‘1‘ C ( 5 pts.) (1) Which weak bond(s) is strengthened by adding some CI'LCHzOH to an aqueous solution containing molecules that exhibit all of the different types of weak bonds? l(..‘1. y)”, mi ( 5 pts.) e) Which weak bond(s) is weakened by lowering the temperature of an aqueous solution containing molecules that exhibit all of the different types of weak bonds? V’ '-\_I 1;: lr 1: “\f- C ( 5 pts.) f) Which weak bond(s) is weakened by adding urea to an aqueous solution contamtng molecules that exhibit all of the difi‘erent types of weak bonds? \J })M_.\\," 3f hutdvx‘w‘lL-W (- iological Sciences 102 Name 3. (continued) (10 pts.) g) How are weak bonds important in enzyme catalysis? Emz,gjm.2 'COV mq \ovxlr bowel w/ Subg+v<ip 4. The change in entropy, AS, is very important in biochelnistIy. ( 5 pts.) a) In what topics in our course does the AS of water play a key role? List all topics. H%&m‘ph 0b“: » (:k‘obs - Wee Bum/(3%)) Q“ ~ (20 pts.) b) Compare and contrast two types of transport mechanisms that rely only on AS to drive the vtransport of solutes across a membrane. QWM Diflfim-iow (4 pts.) c) In chemical reactions, what is the sign (i.e. '+' or '-') of AS’ ? Explain why that is. L—\ We m: is U) ( 6 pts.) d) What do enzymes do to this AS1 term (i.e. do they increase or decrease the magnitude of this g term)? Also explain how they manage to accomplish this change. 'mCKQOSQ waa‘xw-tuéie M 739 SQJCOC€ euvlem ercfifimg‘ . Biological Sciences 102 1-1 Name 5. An enzyme with a K... of 4-0 x 10" M was assayed at the following substrate concentrations: 2.00 x 10" M, 6.66 x 10" M, and 0.050 M. The initial velocity observed at 0.050 M was 3.23 nM min". ( 35 pts.) a) On the graph paper on the next page, make a Lineweaver—Burk plot of the above data. Plot all data points and show the x- and y-axis intercepts. Label your axes (including units). Circle your data points. Make your graph large (i. e. scale your graph to fit most of the graph paper). In order to make the plot, you will have to do several calculations. Show these calculations in the space below. Calculag'ons: Umaw C-OCD-‘D —. m —_ v x7317; 31°) “0'1 3r CDLS—S mane : 7 Cg‘) \bo gm ‘ roe-a __ M 2,5 “we “0M0” V’o '3 “Mama -"I V0 1 3.1% Eugene ‘3 _ 2‘01 Hacker-l & Lb-bb‘#\°’fi\ ._"1\ V0 __ -.~_ \_0% HMO" t Llyko‘“) ,L _\ Vb LS? “0 C31 | .—. 3.11.11) II :05 III IB’I [VD . E l gaueu Le”; ' b Loewa Wag " c ._ 6x \O \‘O% \ 1%\Q *3 .Cich .L 1.- _ VMKX ' 3713* ‘13\ \ \ __ u—‘— 7 ._ .—-.. -_ _2 (a K _ Wag—1 ‘5 x“) Name . Biological Sciences 102 IIIIIIIIIIIII 6 IIIIIIIIIIIII I IIIIIIIIIIIIII IIIII IIII IIIII I I IIIIIIIIIIIIIIIIIII IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII Ill-III.IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII H IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII III-IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII :4 'lIIIIIIIIIIIIIIIIIIIIIIIIIIIIII'IIIIIIIIIIII IIIIIIII-IIIIIIIIIIIIIIIIIIIIIIIII IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII 2 “III-unnununfimnnnnunnuflufiunnnnuflflunnflnnnun co IIIIIIIIIIIIIIIIIIIIIII .1 nflaflnununuflnuflnnnullllllnnnllllnn - u, IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII II.I.n.I..uI.I..!..I...n“an”flunuuunnuflafinnnufl“ w o IIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIIII r...""Hourlmmmmmmfifiuudaufi I.I..IIIII.I....I.II...I.I......IIum.I...II.I - IIIIIIIIII'IIIIIlIIIIIIIIIIIIIIIIIIII-Ill _ III-IIIII II I II I II IIIIIIIIIIIIIIII "m * IIIIIIIIIIInIIII III-I'll-“ II II II Illflflllllllfl 1x I..." nun—n... .nu_._u—_nu. .unuuflquI-III. III-IIIIIIIIIII'II IIIIIIIIII II II 3 IIIIIIIIIIII III I II III II IIIII'II _ II I III IIIIIIIIIIIIIII II I II III II I IIIIIIIIIIIII IIIIIIIIIIIIIIIIIIIIIIIIIIIIIII'IIIIIIIIII - In... I... “- ...- nflunun-fluuflullllll A E II ##IIIIIIIHIIIIII I IIIII III I II I 9 a‘ $\ A\ U\ 6‘ Q 51. 1 \ xi... Lineweaver-Burk plot of the previous data: (continued) plotworddlooklikeifthesamesetoftestmbeswere inhibitor that could only bind to the ES complex was ( 5 pts.) b) On the above graph, indicate what the re-assaycd, but this time a fixed amount of a reversible 3' added to each tube. Label your plot as "+1". \lfimmpek‘xl‘we Biological Sciences 102 Name 6. (5 pts.) Of all of the protein purification techniques available, which one is the “best”? Why? m\f\\-\’V\—5§\\r33 mega pmvéicw‘flo‘fl WC aQ’VQVYWxW-G ‘wl'w 6W4“: '3‘ 7. (5 pts.) How do biochemists judge the purity of an enzyme solution? wdgc achum/r 8. (10 pts.) Predict how the following environmental changes will affect the pK;. of a lysine residue in a protein. Give a brief explanation of your reasoning. a) An aspartic acid residue in the same protein is brought into close proximity. (—3 L-\\ x‘ \Dka Ctr/C N «3 W‘\\\ Cy av) CM is") b) The lysine side chain shifts from the outside of the protein to the inside of the protein. cm 3}, 8x ,5; ~. <V12L..:\(\\( 1. P A z" “ w o gal-*1 V ’ 4c o 3, pkg b(¢ 9S1) (J:\ Tb LL33 -’) \oge PVDAFUVN 9. (10 pts.) To purify enzyme "Z", one needs two different chemical assays to determine the success of each purification step. What are these two assays and why is each important? y . _ anztjme 06353445 ,>,» deiavvmwa L .U- {i}; \3‘ . 7”“ C)“ V’Y’V" {3‘1" ‘31 C \ n 1 .U Ton—o) 10. (15 pts.) Draw the structure of a lipid, in the form that would predominate at pH 7, that satisfies the following criteria: (a) it does not react with ninhydrin or molybdenum blue, (b) it contains a saturated fatty acid, and (c) it contains an unsaturated fatty acid that only plants can make. x t I n- ,, fl. ‘ ' r , ' . “- ‘(Z’( “" V1621? Vb C\»<‘:"‘(“V‘iw F‘s” : ‘ 4’ L\C‘\- \v'd' \ M; mmwwmw \(JhmP‘v’LcthA \MD\€\C 0" \MVWWW 0 “NW mC—D'C \ 1 of 4 BIS 102 Name L, Summer Session II Last First K. Hilt Final Score (134): Equations: pH = pKa + log {[b]![a]} AG = AH — TAS V0: {Vm [5]} f {Km+[S]} F=(q1<t2)f(8r°') l. ( 10 pts.) Draw the structure of a phosphatidylserine molecule that contains one saturated fatty acid and one unsaturated fatty acid. Draw this molecule in the form that would predominate at pH 7. O 0 “AW Cd). “0- C a I .__O___{-““ *1)»; We '- n ‘ ._.D -- PFETCH- C‘r\ 6’61 \ 2.1 .- o coo‘ 2. ( 12 pts.) What is the final pH when 4.00 ml ofSO mM Tris, pH 8.21, is mixed with 1.00 ml of 50 mM HCl? The 1er3 of Tris is 8.21. Show all calculations. 82$ ‘— ‘12, ‘l\ 1‘ \goggé‘ fig —_ \ \0=_l5 (no 2 10 when)" _ (1:63 (qu ) '_ 'ZQNW‘UD\ (5-; ML, L50 mMj ': \ko sax \__:mL L‘BOth'l l0 mmcfl 'v L'D m - D \+—. {5.7.x » \0 29—3—— Q o) lor‘db 3. ( 5 pts.) Derive the steady state rate expression for Kn. Recall that this expression has the rate constants k1, kg, and k4. Show how you derived this expression. ACE‘SCCJX 1 \4,\CE<>‘5 * alcea CE 51“; - tr. 4. ( 5 pts.) 57% pltia of acetic acid in water is 4.76. Would the pK. of acetic acid be higher, lower, or the same if the acetic acid was in a solution of 50% ed1anoU50% water, rather than pure water? Explain. The. ?\’—c\ m'rm‘di meted-’36 DLCONCE aw, mix-c ab: \—r— ., _"'-tj iv". A '96 I: (v {xi-\IQ l.‘,~I-I5L<Q {J I C. 20f4 BIS 402 Name V 5. ( 5 . An irreversible inhibitor could give a series of lines on a Lineweaver-Burk plot that could be co ed with what type of reversible inhibitor (uncompetitive, noncompetitive, or competitiv ? xplain. 6. ( 15 pts.) Compare and contrast analytical SDS-PAGE, native PAGE, and IEF gels. How are they the same or different, in terms of how they are constructed, how they separate proteins, and what they can tell us, separately or combined, about a pure protein? <3§C> ‘ 7003(va 1E? \Iavt‘xrm aadadflwa o’erV (‘50 a 3* “Ck'mq mm \O\)\ CH fiffig u\ Wxa<”.;~‘, ‘ Chg)”. (%Q {t QKZ. e ., \ 2 ~ ‘ t ,K a r .i ‘ . gee 5““ PWN a L :6; 2x ~ sex i «ka w} IE. _, V \ use 52‘“ o \ a, ‘6 ‘ 9 use C§\e,\e(%gv\t am Ag gram 3&0005‘2‘ (“:5 'v\ :9 '5: ‘53”? *r x. at. \m 0&5 3, Va 7. ( 5 pts.) Do you agree with the following statement? “Linus Pauling contributed an important insight into enzyme kinetics. He proposed that enzymes bind tighter to substrates than they do to products.” Explain why you agree or disagree with this statement. vsgt QC 9 m? 9‘3? :13; €-\o:‘2/U§i”\i‘x a“. (5‘ ’ {EVA 6 ‘v‘ Vi) “E V“: \ Q) "" ’Q ’ caWTfif {03 8’3; %\o\N~Q\(% Em 9‘: (it rock. 4.1V 06: Q " , if w 8. ( 8 pts.) Do you agree with the following statement? “Hemoglobin has several noteworthy histidines in its structure. Probably the most important ones are the C-terminal histidines of the B-subunits.” Explain why you agree or disagree with this statement. stss \N 9W '5‘ “AN- i‘u € ext-755* (t *x p‘f‘w w A \v \ ~ v V I t c- C ‘ “\ ‘ ‘ 02 \waa 03' axvrrye “i \ow’xck ' \ . {‘y‘ r“ 1""; , b : r J a“ « \43 var ad to Mar 3&3 0‘ ‘dflQg ' 5 "‘l Com m om 3 of 4 BIS 402 Name 9. ( 5 pts.) An enzyme has a turnover number of 30,000 min“. If this enzyme has a MW of 65,000, what must the initial velocity be, in international Units, of an enzyme reaction that contains 2.0 ug of this pure enzyme? Show all calculations. JU‘ e. I\ I 1‘9: F_“:‘ _\_/in1 \m9\ ' RUDE ‘ EMA Mmcx ’U‘mb“ £1- b‘iooocfi M3 ' meo‘ Elmo ‘— UD: #mok/mlxfl I. a I 3\O%h\9§Mm\ : any) 1m 10. ( 5 pts.) A Michaelis-Menten enzyme is assayed at a [S] = 3 x K.” What percentage of Vm will the observed v0 be? Show all calculations. \anav: \ 1.» ‘-" 'l— Y‘r‘f‘ '2) V __ r H Vt)”: ' ‘7 t f W131“ _' I: ._/".D lam-K2 i: a: mi “K 1 l. 1 ts.) We mentioned the compounds ammonium sulfate and ammonium persulfate in class. What we he context in which each was mentioned? Why is each compound used? How do these compou ccomplish their respective tasks? 12. ( 8 pts.) What is the Current model for how proteins fold? \D "=5\°L0~\ lo “’“CX—OQ 7.)" &omc\.\r\ AMQVWV‘ 5‘3“? NAM». “P " Q .959; was ’5“: “)3” 4 of 4 BIS .102 Name 13‘ ( 6 pts.) What is the current model used to explain the S-shaped curve of oxygen binding to hemogiobin? C) 0—; CD7, 1 \d L E -——-——> a a» as —-== as \"\~u\%"\ "rte/Ne 0A re (ILA owe 02. bCMV‘0\ TO flack dimer. l4. ( 20 pts.) Draw the complete structure of a tetrapcptide that strongly absorbs ultraviolet light, contains the residue that was a nucleophile in the catalytic triad of chymotrypsin, contains a residue that has a guanidino group in it, and contains a residue that needs to be reduced with dithiothreitol in order to prevent disulfide bond formation. Draw the peptide in the form that would predominate at pH 0. UN -=-i Aw“ I W 3?er (Swan/Mi -— OKV% 9.53 “IN-Ag :6 “\‘QIWILIKQ .3, (EQV o E) P C) * _ —“_ — «—“~w—c——‘—~ss-c.—E_H " “3‘4 Cf c C" o x H l D “ (.361, m1 “arm‘s 9"1 x UT‘ \ \y\'4 Ir‘d'i‘Q. I 15. pl 15 pts.) When Watson and Crick looked at Rosalind Franklin’s x-ray crystallographic data of DNA in the early 1950’s, it became obvious to them what the structure of double stranded DNA was and how this structure explained the function of DNA in DNA replication and mRNA transcription. A few years later, when John Kendrew looked at the x-ray crystallographic data of the first globular protein, myoglobin, it was not obvious how proteins were put together or how they function. Years later, we are still a long way away from understanding protein su'ucmreffiinction. Having now had a course in proteins, discuss why understanding protein structurelfimction is much more complex than DNA structurei’firnction. BIS 102 Name Fall, 2002 Last First K. Hilt Final Exam Score (200): 1. You decide to make up 5.00 liters 01°30 mM glycine bufi'er, pH 9.30. ( 20 pts.) a) You have the fully protonated form of glycine, glycineoHCl, FW 111.5, and some 5.00 N NaOH. Indicate how you would make up the above buffer. The pKa’s of glycine are a—COOI—l group 2.34; (I-NH3+ group 9.60. Show all calculations. 5 .C\““J ( 20 pts.) b) You are going to use 10 ml of the above buffer to control the pH of an enzyme catalyzed reaction that produces one H“ for every substrate molecule that reacts. How many I.U.’s of enzyme Z must be present in this 10 ml of buffer if the pH drops to 9.00 after 5 minutes? Assume that the enzyme is operating under V“mm conditions during this 5 min. and that addition of the enzyme and substrates did not change the original buffer concentration and pH. Show all calculations. (. BIS 102 Name 2. A pentapeptide was subjected to the following analysis. a) Treatment with cyanogen bromide yields a tripeptide and free methionine b) Treatment of the intact pentapeptide with trypsin yields a single product. However, treatment of the intact peptide with trypsin and dithiothreitol yields two products. ( 5 pts.) a) What is a possible amino acid sequence for this pentapeptide? (Use the 3—letter code). Sequence: m ‘r I t . "‘~ {a ., axing, ( 5 pts.) b) Is there another possible sequence? Explain. mat - mes ~ My: - Mg ~ we. ( 5 pts.) c) If there is another possible sequence, which endoprotease(s) would you use to determine which sequence was correct? If there is no other possible sequence, disregard this and receive five free points. ' A ~ .‘eeste .w 0:» C. A k-( Q‘x s. N o 9 me x). (it. - e .9. J "T I: ' ~35 Q‘s. {3 (V c\~J\(‘\ *3 2.1"- c‘« J {2’ a 3. ( 5 pts.) How does adding liquid ni ogen to tissue break open cells? ‘OVQMKQ weak law/‘39 4. ( 6 pts.) How is it possible to get two different activity peaks for the same enzyme from an affinity column after eluting the column with a salt gradient? Explain. \Sovsm QS 'gfiqu epWWQ 1m EH 'PW WW 5. ( 8 pts.) An arginine residue in a protein moves away from a glutamate residue in the same protein. What, if anything, will happen to the pKa of the glutamate? Explain. Qua ’t mgkfl (gm k- x «a» w Mm a: «a a . *‘v A O -’> 6&1}. ka‘cw «=5 “x. 9‘40 6. ( 9 pts. Explain how carbonic anhydrase and the anion exchange protein figure into the transport of C02 from the tissues by red blood cells. Use chemical structures to explain the processes involved. Cw com a.» way 0.9.9 eivtwywt ‘erfix <7 queg L02 \ tic-go 1m. am e may Wfip‘s he»; get eu‘i ; e t: ” 9?“ ‘eLO ’3: k we an quhaw o A ‘3) \wrx "’5 EVA \ ov‘ 5 £1531“ ' (x I ' s - \ i i x . 'L «a u ‘7' l. u?) uA’V gagi- Q Q\“ gm“; ' ’3 C0" ‘KDDNN L a»-“‘“”\” (‘3 lips; 4“ V i” (A ’3'“: 4*. V’xqb \ w. he 7. ( 9 pts.) Identify three differentcharaCter‘fst’i‘cs tha'gmake‘forfafigood” buffer: #1 g‘ég; c3955 ’Yb 2H W #2 Y\l\ e! 15—33me Qfi'flégy 'tm‘¥€«€eyeww 8. ( 10 pts.) The names of all of the known allosteric effectors of Hb are (exclude CO): C101+\'\* \_C,\' FEW; Ca (l- BIS 102 Name 9. Palmitate is a sixteen—carbon fatty acid. When palmitate is put into water, the following reaction occurs spontaneously: ( 5 pts.) a) Which weak bond(s) destabilize the formation of the micelle? ll 0 Y\‘\ C ( 5 pts.) b) Which weak bond(s) stabilize the formation of the micelle? Dbl C ( 5 pts.) c) Which weak bond(s) neither stabilize nor destabilize the formation of the micelle? Va A new Nam \ , 3A ~ bo‘fld ( 5 pts.) d) If we lowered the temperature of the system 10 °C, which side of the equilibrium, if any, would be favored? Explain. \l/ T vfimfikfi‘v‘xfi Eutflu‘ufhobfl 10. ( 15 pts.) Rank the following protein purification steps from best to worst, in terms of likelihood of giving the highest purification, compared to the previous step, an ammonium sulfate cut. Best = 1. Worst =5. Put one number in each of the following blanks. Then explain why your best step is the best and your worst step is the worst. Another ammonium sulfate cut 6 Sizing column Ll Preparative IEF gel 1 Affinity column ' Ion exchange column “2 Explanation of why your best step above is the best: Explanation of why your worst step above is the worst: 11. ( 10 pts.) A Michaelis—Menten enzyme is assayed at a substrate concentration of 2.0 x 10'6 M. The resulting initial velocity was 2.4 nmoles P/20 sec. Previous experiments showed that the x-intercept on a Lineweaver—Burk plot for the same-enzyme was “-0.5” on the axis labeled “l/[S] M’1 x 1045 ”. What is the value of Vm, in umoles/min.? Show all calculations. _ vb 7 (i. \ _—b a ‘ C33 -Z 07“© V9 ‘ 1 5K N‘MJ‘ I jifimto (903‘ 'Tm-c " .57 XV *3 km IZX‘ICE’ [[0 (PC \mm r.‘ , V «x CZXVDJ’3 ‘3 ’7‘Zx\o 0‘ - Nmfimr \jwvv ‘— 7. Z >UU \1 “ACYISWCEQ ., . “ “ (2.0Mbe )r¥2+\b' was ‘ L 3 V0” BIS 102 Name 12. ( 6 pts.) What stabilizes the formation of an (it—helix? If there is more than one thing, then list them all. l/Mcimflvo‘o'w, H Mend 13. ( 20 pts.) You have been a given a newly discovered plant from the South American jungles. How will you determine the lipid and fatty acid composition of the leaves? Indicate all materials, equipment, and procedures necessary to identify and quantify each. ghee u V —>.: 9m 1m Stag: w} worm *1 for) we” a flei'ik‘VV‘ *3 “3 are“ " ' ' 1 y «M. A _, Q » w l A. A 3",.- \ A. . 7 “3C\(t “(A C v~ \ é‘ t: manicuzi v A»), A Cam/“55‘7" C 5'1“ “' 8° ‘v' " U1. f“ "1“. wCMA\ \QQJ.’ “:5 to“); (K g}; (1‘an C. n R.» u t" y )) L;\O‘\& J I 30 TLC m war:qu U“\‘V\°V°"‘ . Odéx 904‘) 34‘ 9\0‘)\"e aevpt’q “A (OKAWD “Ck ‘1 vim * SJ‘flde‘Cw/OW “4333/?ch W/HinvijN/‘lni SC\Y\&V\\\O\*1\QV\ kMAOV Tu Jdctgvmw X QM» W’k'VxW' \ P Netti 2’39 WVMWSJV w 90 WP Ckwwmo’“fl’awml TNW. 9m Vacttoacilwl‘“ V 14. ( 12 pts.) Do you expect the plant leaves in question #13 above to contain more: i) stearic acid or palnnfic acid? (circle the correct answer) ii) oleic acid om (circle the correct answer) Explanations: H "t V106 ms re. : \oonA“; )3 €‘f3‘36‘fl+ K at u" 15. ( 15 pts.) After many purification steps, a student thinks that their enzyme solution is pure. They take separate small aliquots (samples) and run three different analytical gels —— a Native PAGE gel, an IEF PAGE gel, and 3 SDS-PAGE gel. All three gels are stained and developed with Coomassie Blue, a general protein stain. The native PAGE gel yields one band. The IEF gel yields two bands. The SDS-PAGE gel yields three bands. Do you think that this enzyme is pure? If you do, then explain how all of the results are consistent with this conclusion. If you do not think that the protein is pure, then explain how all of the results are consistent with that conclusion. 5‘08 Amun‘w vzmptmws w Vail“? NoéQre, A; '2, aflfl/ksxmex Manx “vfioxoe SCINOQ Size 7 r . v , - . , \\\ . . ‘. x 7 ‘ r7 : X (a x. . ,I \\ “‘2' f“ (‘3 w \ a V .E . \ NU)‘ QXGWQ v )Ic 1" V a 5 Q Adv” ...
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This note was uploaded on 01/31/2012 for the course BIS 102 taught by Professor Hilt during the Winter '08 term at UC Davis.

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BIS 102-Hilt-Final - Biological Sciences 102 Name Winter,...

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