TY - JOUR
T1 - Characterization of the catalytic pathway for D-serine dehydratase. Evidence for variation of the rate-determining step with substrate structure.
AU - Federiuk, C. S.
AU - Bayer, R.
AU - Shafer, J. A.
N1 - Copyright:
Medline is the source for the citation and abstract of this record.
PY - 1983/5/10
Y1 - 1983/5/10
N2 - Steady state kinetic parameters (kcat and Km) which were determined for the D-serine dehydratase-catalyzed decomposition of the isomers of serine and threonine at pH 5.7, 7.8, and 8.9 indicated that the enzyme exhibited considerable kinetic specificity. At pH 7.8 and 25 degrees C, the specificity constants (kcat/Km) were as follows: D-serine (3.4 X 10(5) M-1 s-1), D-threonine (2.9 X 10(4) M-1 s-1), D-allothreonine (7.5 X 10(3) M-1 s-1), and L-serine (38 M-1 s-1). Substrate C-2 deuterium atom isotope effects on the steady state kinetic parameters disclosed that the rate-determining step in the catalytic pathway varied with substrate structure and pH. Removal of the C-2 hydrogen atom of the substrate was shown to be fully rate-determining with L-serine and partially rate-determining with D-allothreonine as substrate at pH 7.8. Stopped flow measurements of absorbance and fluorescence were used to characterize intermediates in the catalytic pathway. These measurements indicated that D-serine, in addition to being the best substrate, was processed faster than the other substrates through steps in the catalytic pathway which were not rate-controlling. Kinetic evidence also was obtained which indicated that the base which accepts the proton from the C-2 carbon atom of the substrate must be aprotic. Thus, the catalytic site of D-serine dehydratase should contain a basic group in addition to the active site lysyl residue.
AB - Steady state kinetic parameters (kcat and Km) which were determined for the D-serine dehydratase-catalyzed decomposition of the isomers of serine and threonine at pH 5.7, 7.8, and 8.9 indicated that the enzyme exhibited considerable kinetic specificity. At pH 7.8 and 25 degrees C, the specificity constants (kcat/Km) were as follows: D-serine (3.4 X 10(5) M-1 s-1), D-threonine (2.9 X 10(4) M-1 s-1), D-allothreonine (7.5 X 10(3) M-1 s-1), and L-serine (38 M-1 s-1). Substrate C-2 deuterium atom isotope effects on the steady state kinetic parameters disclosed that the rate-determining step in the catalytic pathway varied with substrate structure and pH. Removal of the C-2 hydrogen atom of the substrate was shown to be fully rate-determining with L-serine and partially rate-determining with D-allothreonine as substrate at pH 7.8. Stopped flow measurements of absorbance and fluorescence were used to characterize intermediates in the catalytic pathway. These measurements indicated that D-serine, in addition to being the best substrate, was processed faster than the other substrates through steps in the catalytic pathway which were not rate-controlling. Kinetic evidence also was obtained which indicated that the base which accepts the proton from the C-2 carbon atom of the substrate must be aprotic. Thus, the catalytic site of D-serine dehydratase should contain a basic group in addition to the active site lysyl residue.
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M3 - Article
C2 - 6406501
AN - SCOPUS:0021099428
SN - 0021-9258
VL - 258
SP - 5379
EP - 5385
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 9
ER -