Passage
Leukocyte common antigen (LCA) enzymes play several important developmental roles in both vertebrates and invertebrates. They regulate numerous cellular processes by catalyzing Reaction 1 in target proteins at specific developmental stages. Consequently, LCA enzyme expression and activity are tightly controlled in various cell types.
Reaction 1The Src protein is critical for cell differentiation and motility, and is activated by LCA enzyme activity. Two LCA enzymes, known as LAR and CD45, were purified and incubated with the phosphopeptide src527 (Figure 1) , derived from Src.
Figure 1 Structure of the src527 peptide used as a substrate in LAR and CD45 assaysThe rate of Reaction 1 was measured at various concentrations of src527 in the presence of either LAR or CD45, and kinetic parameters were determined (Table 1) .Table 1 Kinetic Parameters of LAR and CD45
Compound 1 was synthesized and tested for its effectiveness as an inhibitor of LCA enzymes.
The study in Table 1 was repeated in the presence of Compound 1, which was found to significantly reduce the activities of both LAR and CD45 against the src527 peptide. Compound 1 was subsequently used in assays to examine the in vivo function of LCA enzymes.
Adapted from Cho H, Krishnaraj R, Itoh M, et al. Substrate specificities of catalytic fragments of protein tyrosine phosphatases (HPTP beta, LAR, and CD45) toward phosphotyrosylpeptide substrates and thiophosphotyrosylated peptides as inhibitors. Protein Sci. 1993;2(6) :977-84.
-Based on Table 1, the turnover number (k_cat) of CD45 exceeds that of LAR by what factor?

Question

Passage
Leukocyte common antigen (LCA) enzymes play several important developmental roles in both vertebrates and invertebrates. They regulate numerous cellular processes by catalyzing Reaction 1 in target proteins at specific developmental stages. Consequently, LCA enzyme expression and activity are tightly controlled in various cell types.

Reaction 1The Src protein is critical for cell differentiation and motility, and is activated by LCA enzyme activity. Two LCA enzymes, known as LAR and CD45, were purified and incubated with the phosphopeptide src527 (Figure 1) , derived from Src.

Figure 1 Structure of the src527 peptide used as a substrate in LAR and CD45 assaysThe rate of Reaction 1 was measured at various concentrations of src527 in the presence of either LAR or CD45, and kinetic parameters were determined (Table 1) .Table 1 Kinetic Parameters of LAR and CD45

Compound 1 was synthesized and tested for its effectiveness as an inhibitor of LCA enzymes.

The study in Table 1 was repeated in the presence of Compound 1, which was found to significantly reduce the activities of both LAR and CD45 against the src527 peptide. Compound 1 was subsequently used in assays to examine the in vivo function of LCA enzymes.
Adapted from Cho H, Krishnaraj R, Itoh M, et al. Substrate specificities of catalytic fragments of protein tyrosine phosphatases (HPTP beta, LAR, and CD45) toward phosphotyrosylpeptide substrates and thiophosphotyrosylated peptides as inhibitors. Protein Sci. 1993;2(6) :977-84.
-Based on Table 1, the turnover number (k_cat) of CD45 exceeds that of LAR by what factor?

Quizplus · Accepted Answer

11ecba2d_0ea8_54d4_a3bf_41b6a197e3fc_MD0008_00 The turnover number (k_cat) of an enzymatic reaction is the number of reactions catalyzed per second per enzyme under saturating conditions (ie, when V_max is achieved). It is useful for comparing enzymes when substrate concentration is high and can be calculated by dividing V_max by the total enzyme concentration [E]. Table 1 does not show the turnover numbers for either CD45 or LAR, but each can be calculated from the given V_max and [E]: For CD45, V_max = 2.8 μM/s and [E] = 0.01 μM. Therefore, 11ecba2d_0ea8_7be5_a3bf_9d07ecf71024_MD0008_00 kcat=2.8 µM/s0.01 µM=280 s-1. For LAR, V_max = 2.0 μM/s and [E] = 0.05 μM. Therefore, 11ecba2d_0ea8_7be6_a3bf_2d72c64250ef_MD0008_00 kcat=2.0 µM/s0.05 µM=40 s-1. The ratio of the two k_cat values is 11ecba2d_0ea8_a2f7_a3bf_b9fbe17bd526_MD0008_00 28040=7. Therefore, the turnover number of CD45 exceeds that of LAR by a factor of 7. (Choice A) The ratio of V_max of CD45 to V_max of LAR is approximately 3:2, so V_max of CD45 is approximately 1.5 times greater than that of LAR. However, CD45 is also present at a concentration [E] that is 5 times smaller than that of LAR. Because k_cat = V_max/[E], the enzyme concentrations must be taken into account. (Choice B) The ratio of LAR K_m to CD45 K_m is approximately 2:1, but this is not the ratio of turnover numbers (k_cat). (Choice C) The concentration of LAR exceeds that of CD45 by a factor of 5, but enzyme concentration [E] is not the turnover number. Educational objective: The turnover number k_cat of an enzyme-substrate system represents the number of reactions catalyzed per second per enzyme under saturating conditions. It is calculated as the V_max divided by the total enzyme concentration [E]. It can be used to compare enzymes at high substrate concentrations.