Crystal structure of the complex of carboxypeptidase A with a strongly bound phosphonate in a new crystalline form: Comparison with structures of other complexes
- Harvard Univ., Cambridge, MA (USA)
O-(((1R)-((N-(Phenylmethoxycarbonyl)-L-alanyl)amino)ethyl)hydroxyphosphinyl)-L-3-phenyllacetate (ZZA{sup P}(O)F), an analogue of (benzyloxycarbonyl)-Ala-Ala-Phe or (benzyloxycarbonyl)-Ala-Ala-phenyllactate, binds to carboxypeptidase A with great affinity. Similar phosphonates have been shown to be transition-state analogues of the CPA-catalyzed hydrolysis. In the present study, the structure of the complex of phosphonate with carboxypeptidase A has been determined by X-ray crystallography to a resolution of 2.0 {angstrom}. The structure of the complex was solved by molecular replacement. Refinement of the structure against 20,776 unique reflections between 10.0 and 2.0 {angstrom} yields a crystallographic residual of 0.193, including 140 water molecules. The two phosphinyl oxygens of the inhibitor bind to the active-site zinc at 2.2 {angstrom} on the electrophilic (Arg-127) side and 3.1 {angstrom} on the nucleophilic (Glu-270) side. Various features of the binding mode of this phosphonate inhibitor are consistent with the hypothesis that carboxypeptidase A catalyzed hydrolysis proceeds through a general-base mechanism in which the carbonyl carbon of the substrate is attached by Zn-hydroxyl (or Zn-water). This complex structure is compared with previous structures of carboxypeptidase A, including the complexes with the potato inhibitor, a hydrated keto methylene substrate analogue, and a phosphonamidate inhibitor. Comparisons are also made with the complexes of thermolysin with some phosphonamidate inhibitors.
- OSTI ID:
- 5449890
- Journal Information:
- Biochemistry; (United States), Vol. 29:23; ISSN 0006-2960
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
CARBOXYPEPTIDASES
X-RAY DIFFRACTION
PHOSPHONIC ACID ESTERS
CRYSTALLOGRAPHY
ENZYMATIC HYDROLYSIS
ENZYME INHIBITORS
CHEMICAL REACTIONS
COHERENT SCATTERING
DECOMPOSITION
DIFFRACTION
ENZYMES
ESTERS
HYDROLASES
HYDROLYSIS
LYSIS
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
PEPTIDE HYDROLASES
SCATTERING
SOLVOLYSIS
550602* - Medicine- External Radiation in Diagnostics- (1980-)