(c) ( To identify the critical enzyme resi- due:substrate interaction and determinant of sub- strate specificity, steady-state kinetic studies have D(-4) been carried out by site-specific mutagenesis of the enzyme and by systematically changing amino acid residues in the 11-amino acid phosphotyrosine con- taining substrate that simulate residues 988 - 998 of the epidermal growth factor receptor (EGFR988-998). The table below compares the kinetic parameters of the wild type enzyme with phosphotyrosine pep- tides in which amino acid residues are substituted. In the adjacent diagram the structural interactions between active site residues are illustrated for the first 6 residues of the substrate. Identify which sub- strate residue is most sensitive to substitution and justify your conclusion on the basis of the kinetic parameters. Is the decrease in kcat/ KM due to loss of catalytic reactivity or due to suboptimal position- ing of the phospho-Tyr residue in the active site? Substrate DADEPYLIPQQG DADAPYLIPQQG DAAEPYLIPQQG AAAAPYLIPQQ G kcat 44.6 1.8 39.8 ± 0.32 35.3 ± 0.22 34.7 ± 0.25 A(-3) D(-2) Tyr46 Ser216 Arg47 E(-1) Asp181 PTP1B Km μM 3.9 ± 0.9 13.7 ± 0.46 52.7 ± 0.7 6.6 ± 0.22 Y(0) Asp48 L(+1) Phe 182 Gln262 kcat/Km 10-7 X (S¹M¹) 1.1 ± 0.25 0.29 ± 0.01 0.53 ± 0.02 0.066 0.001
(c) ( To identify the critical enzyme resi- due:substrate interaction and determinant of sub- strate specificity, steady-state kinetic studies have D(-4) been carried out by site-specific mutagenesis of the enzyme and by systematically changing amino acid residues in the 11-amino acid phosphotyrosine con- taining substrate that simulate residues 988 - 998 of the epidermal growth factor receptor (EGFR988-998). The table below compares the kinetic parameters of the wild type enzyme with phosphotyrosine pep- tides in which amino acid residues are substituted. In the adjacent diagram the structural interactions between active site residues are illustrated for the first 6 residues of the substrate. Identify which sub- strate residue is most sensitive to substitution and justify your conclusion on the basis of the kinetic parameters. Is the decrease in kcat/ KM due to loss of catalytic reactivity or due to suboptimal position- ing of the phospho-Tyr residue in the active site? Substrate DADEPYLIPQQG DADAPYLIPQQG DAAEPYLIPQQG AAAAPYLIPQQ G kcat 44.6 1.8 39.8 ± 0.32 35.3 ± 0.22 34.7 ± 0.25 A(-3) D(-2) Tyr46 Ser216 Arg47 E(-1) Asp181 PTP1B Km μM 3.9 ± 0.9 13.7 ± 0.46 52.7 ± 0.7 6.6 ± 0.22 Y(0) Asp48 L(+1) Phe 182 Gln262 kcat/Km 10-7 X (S¹M¹) 1.1 ± 0.25 0.29 ± 0.01 0.53 ± 0.02 0.066 0.001
Biochemistry
9th Edition
ISBN:9781319114671
Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Chapter1: Biochemistry: An Evolving Science
Section: Chapter Questions
Problem 1P
Related questions
Question
Expert Solution
This question has been solved!
Explore an expertly crafted, step-by-step solution for a thorough understanding of key concepts.
Step by step
Solved in 2 steps
Recommended textbooks for you
Biochemistry
Biochemistry
ISBN:
9781319114671
Author:
Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Publisher:
W. H. Freeman
Lehninger Principles of Biochemistry
Biochemistry
ISBN:
9781464126116
Author:
David L. Nelson, Michael M. Cox
Publisher:
W. H. Freeman
Fundamentals of Biochemistry: Life at the Molecul…
Biochemistry
ISBN:
9781118918401
Author:
Donald Voet, Judith G. Voet, Charlotte W. Pratt
Publisher:
WILEY
Biochemistry
Biochemistry
ISBN:
9781319114671
Author:
Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Publisher:
W. H. Freeman
Lehninger Principles of Biochemistry
Biochemistry
ISBN:
9781464126116
Author:
David L. Nelson, Michael M. Cox
Publisher:
W. H. Freeman
Fundamentals of Biochemistry: Life at the Molecul…
Biochemistry
ISBN:
9781118918401
Author:
Donald Voet, Judith G. Voet, Charlotte W. Pratt
Publisher:
WILEY
Biochemistry
Biochemistry
ISBN:
9781305961135
Author:
Mary K. Campbell, Shawn O. Farrell, Owen M. McDougal
Publisher:
Cengage Learning
Biochemistry
Biochemistry
ISBN:
9781305577206
Author:
Reginald H. Garrett, Charles M. Grisham
Publisher:
Cengage Learning
Fundamentals of General, Organic, and Biological …
Biochemistry
ISBN:
9780134015187
Author:
John E. McMurry, David S. Ballantine, Carl A. Hoeger, Virginia E. Peterson
Publisher:
PEARSON