Question

only need answer for question d and e. thanks

2. Captopril was the first of a class of blood pressure lowering drugs that work by reversibly inhibiting Angiotension-converting enzyme (ACE). Later to the market was Lisinopril. This class of drugs is the fifth most prescribed, approaching 200 million prescriptions per year in the US alone. ACE inhibitors work by preventing the proteolytic cleavage of angiotensin I by ACE to create its active form angiotensin II, which binds to receptors in the walls of blood vessels to initiate their relaxation, reducing hypertension The data in the table below represent a kinetic analysis of human ACE activity in the absence and presence of 50 nM Captopril, 50 nM Lisinopril, and a fraction of blood serum that passed through a 100 kD pore-size filter but was retained by a 50 kD pore-sized filter during ultrafiltration S ELE . - = _ ACE activity was followed photometrically using the artificial substrate FAPGG. (N-13-02- furyl)acryloyl)-L-phenylalanylglycylglycine, which absorbs at 340 nm Cleavage by ACE eliminates the absorbance. Reaction mixtures contained FAPGG, 300 mM NaCl, 10 uM ZnCl2, 25 mM HEPES, pH 8.2, plus ACE. Incubation was at 37° C. SSSSSSS ACE inhibition kinetics 3 3333 [FAAPGG] V nmol/min2333333 M no inhibitor + captopril + lisinopril +50-100 kD serum 125 0.95 0.13 0. 047 0 .42 167 1.05 0.19 0.061 0.53 L 250 1.54 0.28 0.092 0.65 500 1. 9 50. 460 .18 0.91 750 2 0.61 0. 26 3 951.08 a. Use the above data and Excel to make two fully labelled diagnostic Lineweaver-Burk plots: one with the no inhibitor, captopril, and lisinopril data, the other with the no inhibitor and 50-100 kD serum data. Display trendlines and trendline equations on the plots. In using Excel to make the needed calculations, do not round off the calculations in Excel. b. What are Km and Vmax for the ACE? c. What types of inhibitor are captopril, lisinopril, and in the 50-100 kD inhibitor in the serum? Justify your answers. d. What are the Ki's for captopril and lisinopril? e. Which of the drugs is more efficient and by what factor, i.e., how many fold stronger? Use the ratio Ki captopril/Ki Lisinopril to determine relative strength of binding to ACE

Answer 1

157 Vo (nmollonin) um no inhibitor + Captopril + disinopril 125 0.95 0.13 0.047 167 1.05 0.19 0-061 250 1.54 0.28 0.092 500 1.9 0.46 0.18 750 0.61 0.26 Fore captopril inhibitor : If we calculate the km for ACE in the absence of any inhibitor, then km = 212-84 Umax = 2:56 . In the presence of captopríl , km = 2117-64 and Vomax =2-33. The following results are calculated by equation V=Vmax [5] Km+ [5] 94 is clearly observed that Km increases in presence of captopril inhibitor, but Umax remains almost constant. This is the case of competitive inhibetion . In case of competitive inhibition ,. V = Vmax [5] a km + [5] To calculate de Vo = Rm+ [5] Vmax [s] a km + [5] . where Vo = Initial velocity in the absence of inhibitor vó - Onitial velocity ú the presence of inhibitor

7 0.95 Vmax [s a1284 + 125 Vmax [5] 2117.640 +1a5 0.13 - 7.30 - 207.64 + 1a5 337.84 =>d=11] Now, d = 1 + [1] Riº k 139 1:1+1+50 => 0.1 = 50 = ki = 500 Thus Ki for captopril is 500 Tore lisinopril inhibitor of we calculate the km for ACE in the absence of any inhibitor, then km = 212.84 , Vmax=2.56 In the presence of kasinopril, km = 7261.36 Vmax = 2.77. In this case also, Vmax remains almost constant but km Encreases, this is the case of competitive inhibition . Now for calculation of d, 0.95 Vmax [3] 212.84 + 125 0.047 Vmax [s] 1 7261.360 t1a5 7261.360 + 125 7 20:21 - 12=0.92 337.84 Nowo, 0.92 = 1 +50 = - 0.08 = 5D > Ki-6as Thus ki for lisinopril as - 625

The inhibitor constant, ki is an indication of now potent is an inhibitor. It is the concentration required to produce half maximum inhibitor The smaller the ki, the greater the binding affinity and smaller amount of medication is required in order to inhibit the activity of the enzyme So, by comparing the ki of captopril and lisinopril, ki of lisinopril <ki of captopril - 625 L 500. So, lisinoporil doug is more efficient. ki captopril = 500 = 0.8 ki lisinopril - 625 Lisinopaul is stronger than captopril by 0.8 times.