Exam_1_s08_key - Spring'08 CH370 CH387D Name l I Hackert Exam I UTeID Given c = 3.0 x 10‘" cm/sec k = 1.38x10'23J/K h = 6.63 x 10'3“ J-sec

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Unformatted text preview: Spring '08 CH370/ CH387D Name l: I Hackert Exam I UTeID Given: c = 3.0 x 10‘" cm/sec; k = 1.38x10'23J/K; h = 6.63 x 10'3“ J-sec; N0 = 6.02 x 1023 /m 1. Identify each amino acid by its three m one letter codes (e. g. Ala / A, etc. ) in the blanks above (16) the structures. A) Am [AZ B) Len [L C) Val [V ‘D) The— + - _ .. H3N—ClH—COO H3N+_C\H_Coo' H3Ni-C‘H—COO H3Ni—clH—Coo FHZ :CHz £13 H3C/C13 2 \‘0 CH3 E) ii M F) L I G) H) + - + — + - I, I‘lg:> H3N—CIH—COO H3N—CH—COO H3N—C‘H—COO H3N+_CIH_COQ' a /CH2 /(J:H2 /CH2 CH2 H2C\ H2C\ H2C\ __ S CH2 C\ + \o HN NH H3C/ H2C\/ H2N’ \/' NH3+ 1) M J) M K) L) M + - + - + - Hzn—CIH—Coo H3N—C\}(I:I—_ICOO H3NLCIH_COO- H3N—ClH—COO HZC CH2 / 2 CH\ \CHZ H2C\ Hs/CH2 H c—CH2 CH3 coo 3 M) [152 N) A35: K 0) Z t P) H3NLCIH—coo' HsN— I ‘C00 HgN—CIH—COO' H3N‘i—c11—Coo' CH I CH2 HzC: 2 CH2 CH2 / K /CH2 HN\ \ NH ,C=NH2 HO H2N 2. The dissomahon of an ollgomerlc protein into its subunits IS shown below. If it 18 observed that this (5253/ protein tends to “fall apart” when stored in the cold: What does that tell you about the value of AS for the process shown? Explain why this might be the case from your understanding of protein folding. 6%:30 Acgtbl-\~'FAS (4) ‘t’or (Kw 0L5 WYT-ll’ew be ® RM NM? [3% (a Cold . x w W “wimmwsa {16B 1331,64: c.ka M13, Q’OY elves. fl“ dtssoe, fikQGSJz/S l’Vvdwv\vx0\o\~c, 660sz => L355,“ ® 2. Consider the following oligopeptide: 9K~104 IQ to 8’ (a .2 no tgfiaE-R-A-Y—C-L—V-T-H-F—Q\0H ‘ I (‘l fl: clear a) How many titratable protons are there in this oligopeptide under normal conditions? 2 to # (2) one (OK I'll WorKguw b) What is the net charge on this oligopeptide at very high pH (~pH = 14)? " 51 f \P (3) Q '4 to 8’ IO ID 1.2 “MA QM" "M ~ +3—a+a——r+t—-=20—=v~L—-+-°2~a~3—+-4 1. ) What is the approximate pI for this oligopeptide? 2 (4) d) Starting at the top with the first a.a. of the oligopeptide, use the wheel diagram on right and your understanding of protein secondary structure to predict whether this oligopeptide could form an arnphipathic alpha helix? 0) (3) Check one: 1/ Yes No If yes —Draw a circle on the diagram around those a.a. V residues that would most likely be packed against the interior of the protein. 3. Reco nition of Terms: Match each of terms with the phrase that best describes it. (6) _ Blosum62 a) homologous sequences within a single species that arose by gene duplication _T_ BLAST b) multiple sequence alignments V \0 _e__ orthologous c) independent folding unit with a subunit (‘ a“ _3_ quaternary structure (1) covalent structure L PAM e) homologous sequences in different species that arose from a common ancestral gene during speciation _Q._ domain f) substitution matrix g) arrangement of subunits h) Dayhoff unit to quantify amount of evolutionary change i) sequence comparison algorithm / search sequence databases 4. Consider the glycyl residue below. Label the bond torsion angles phi (p and psi w , and determine what those values are for this residue: (p = (g H o ; w = O o (4) U) (I) \ 5< QXNN (0 O ‘1 H q,» « / O N m H 5. You have the following amino acid sequence from an enzyme that you believe to be a new decarboxylase for converting many amino acids to energy in bacteria. S-Y-V-N—V-K-L-P-H—R-S- D-E-Q-L-K-N—L-V- T—D—A-V-E-P-Y. You search for homologues to find out more about the properties of this protein and its relatives. You use your web browser to go to the well known NCBI bioinformatics web site that provides ready access to many sequence databases. You search for homologues using the BLASTP routine and obtain the following results. E-score l. gi[lO77879] HUMAN PROTEIN 385 2e-49 2. gi[2456879] HYPOTHETICAL PROTEIN gallus 127 463-11 3. gi[8270428] UNKNOWN PROTEIN FROM 2D—PAGE 79 7e—3 4. gi[9667419] Yb42a protein [Helicobacter pylori] . 25 6.95 Q 3 a) What can you conclude from the search results? Are there any homologues? (fl 4)“ x (2) Yes " «(Ml-k m ‘Qmi owe, hOMO\030M/S. b) Aligning the amino acid sequences gives the following results: 5 10 15 20 25 1 A. l . gi [1077879] S-Y-V—N-V L—P H R-S*D—E-Q-L-K-N-L-V-T A-V-E-P-Y 2 . gi [245687 9] A—F-L—-Q-A L—G H L-V—N-Q—Q-L-K-N—L-V—T A-V—E-P-F 3 . gi [8270428] A-Y-V—N-V A-P H Q-S-D-Q—L-K—D-I-V-R-E T-Q—A-V—E Place an (*) above those residues most likely to be involved in the binding and catalysis for this new form of a generic decarboxylase for amino acids . Why, justify your choices? ink-Qt wah NCL. residues, Rte-Yin binds OMCLt kg)“: x M3 “in duMhax 17%“: a; r‘m‘l’ 000' a a ' —— 1 in O \ “‘ T - a If goetxk—MIKM YM‘ _ g M.© u? \ N “NHL? %a.&n V We“ H q _- MCL sum as [vibe cum ‘l’w /d_ev~o\r m moi /LJCLSL ed. 6. e discussed four general propertles of p ems whose d1 ferences can b explmted to separate and purify proteins. Fill in the blanks with either the missing “property” or representative “separation technique” based on that property. (3) (property) (representative method) A) _Size - _‘ifig ikdgj‘m Q‘M‘Uw» / cifixitlsfé B) 50 " 4V - _ammonium sulfate cuts 0) C) tkowé a - _DEAE column D) Briefly describe a major advance Biotechnology has provided us to take advantage of “Affinity” as a property to isolate any targeted protein of interest. (4) 6fo+£C\l\V\o\o \Nlé oviAeQA chm?“ Makers MA fix 6990“ W ohs as} M Your; Tmodwmgfl'e‘g‘mivii3 + s mwtw 5 010 ‘ngueud Votith (a. . tits ~CaX ‘l'a. , Alec cm %hufi Iii/\th ¢O\w~\ws (N3 +Qoi'3 To big m fiffiquayl pro {fix‘ mesh ofi’ M \AOJY-CM$\ ml m eh‘i't s Aim \ANL on Kmihs oLL AAA‘.M+. T cm W rumon lay) RWMM-go‘te Q’W‘hhfzkt «A 7. For the 5—step enzyme purification shown, answer the questions below: Step Protein Volume Total (mg/mL) (mL) Activity Activity (units) —- 1’. Crude extract . 475 13,755 18’ 12 8 27.3 13,400 ' 1 6 . mu 3% 7 I l «3 m 3 11. .1 I Q l 0?. to .m 3382) a) Complete the last column in the Table above. 3.11% __._ .(o Ll b 7 7 (2) b) What is the overall percentage “yield” for this purification scheme? § ' 0 2 Q)“ (2) 0) Which step (#) of the purification produced the largest % increase in specific activity? ’31 I \ (2) d) Which step (#) of the purification produced the smallest % increase in specific activity? 5 i luv” 8. Consider a “gel filtration” column that is 150 cm in length and 2.50 cm in diameter. It is packed with spherical beads that are 0.13 mm in diameter with a V0 that is 33% of Vtot. The column is calibrated with trypsin inhibitor (~21.5 kD) and B-galactosidase (~116 kD) which gave Ve N 0 values of 2.63 and 1.44, respectively. An unknown protein has an elution volume of 411 mL a). Calculate the partition coefficient for the unknown protein. \im - ivrrr‘ 2 736m»? : 739m. 3 Vo‘-35“’nf “5N— , = w 1 57 a V” liqS _+(o c rru‘i “SK 9,193 a 9. A protein has a m lar extinction coefficient of 137,450 M'lcm'1 at 280 nm. A sample in a standard 0.50 cm euvette was found to have a T of 42% at a wavelength of 280 nm. The molar extinction coefficients of tryptophan (a = 5.6 x 103 M'lcm'l) and tyrosine (a = 1.4 x 103 M'lcm'l) at 280 nm are as given and the molecular weight of the protein is 149,000. a) What is the absorbance for this sample protein solution? (3) A = - MALT) = “MAUIQW = 0.370 b) Calculate the E(1%) extinction coefficient for this protein at 280 nm. -17 I? t l: °»M\UI= [0%) :7 E ‘= IOUBHSW : 7.34.,» I I ( I "l i. 0731) @4L3(““> c) Calculate the concentration of this protein solution in mg/mL. I 7. (4) A ‘5 E ' C. * 52. 0.377 = Mail... (micso at) =7 Q = 0.08.; My. UK Mr” . . 5.x 3? = . M ‘9‘ C’M/izjv‘l'rw 4 > O 83 fig“ ‘ I . (4) jyyo WHATS 9. Consider a fluorescent molecule attached to a small peptide. This fluorescent molecule can be excited at 480 nm and emits at 520nm. N ole ‘2 C y) a) Draw a diagram with labels to explain why there is a red shift between the excitation wavelength and the fluorescence emission spectrum. @Excilzl'mm— deck.» aLsarlos (4) watt muwkéfia .3, o M excited s a 9, R7” @ © Fluoresuwu, 7\F>3E dm‘i’o low M b) Name four fluorescence properties that can be monitored for the sign% om this chromophore. 1. 2. i y 6 (4) 3. mm‘ 4. ' v "% ma , 3 9M3 TEMA'QV’ b) Qumlmwb c) Now assume that the peptide binds to a larger protein in such a manner that buries the chromophore. What fluorescence ro erties of the chromo hore are likel to c e and how can we «use that change in behavior to measure the binding of the small peptide to the protein? (You may use diagrams and/or words to explain your answers.) algal; “Waresz Vast/Seas N +0 QKV:VW\M“: (6) émk «,5 WM Mflv‘ M36qu ‘elc. _____.. M t 6 nglfi I) SMIQ‘Y M WW“ V'olloxu Slm Md MJL OT W55 To MmT+ov 070 5E) lDFAAF-M) Takinb VMLLL V5.1L3. *QX Améc‘l'm '5le WM filewegl/xore will Glam +0 views v IMMA’O“ c“ “M 5L Tm MTSOJVWEA To Acfirmtnt 7o \OSAAWQ VSI L . 10. Consider a FRET experiment with condition ch that two chromophores have a R0 4.0 A. What is the separation for these two chromophores if the efficiency of energy transfer is 67%? l g} m_ 1 ° ~l+v<“’ ~ all)" t' r/ a D maQ‘MV + a?) / ‘(01+’G7xb:)ra ~Q {10¢ MEVWW K: «luv Kb: ‘° X > X? R: osmoRo: 30,pr ' ‘1 (Please Sign your name on the back of your exam in a manner tZaz‘ you can recognize it when it is returned.) 5 ...
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This note was uploaded on 03/23/2008 for the course CH 370 taught by Professor Hackert during the Spring '08 term at University of Texas at Austin.

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Exam_1_s08_key - Spring'08 CH370 CH387D Name l I Hackert Exam I UTeID Given c = 3.0 x 10‘" cm/sec k = 1.38x10'23J/K h = 6.63 x 10'3“ J-sec

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