Rate determining step in phospholipase A/sub 2/ mechanism: /sup 18/O isotope exchange determined by /sup 13/C NMR

Conference · Thu May 01 04:00:00 EDT 1986 · Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States)

OSTI ID:5283397

Fanni, T; Lombardo, D; Plueckthun, A; Dennis, E A

Pancreatic and venom phospholipase A/sub 2/ display a marked preference for micellar substrates and act poorly on monomeric substrates. The authors have now examined the merit of the proposal that the product-release step is slow, but is accelerated when the enzyme acts on aggregated phospholipids. Measurements of H/sub 2/ /sup 18/O isotope exchange into specifically-labeled substrate was used to obtain information on the rate-limiting step in the enzyme action. A novel technique of distinguishing /sup 18/O incorporation by /sup 13/C-/sup 18/O vs /sup 13/C-/sup 16/O chemical shift differences at 126 MHz for /sup 13/C NMR was employed. The enzymatic hydrolysis of a micellar phosphatidylcholine analogue of platelet activating factor, 1-alkyl, 2 (1-/sup 13/C)-lauroyl-sn-glycero-3-phosphoryl-choline proceeds by O-acyl cleavage of the sn-2 ester bond. The reaction was examined for the possibility of simultaneous /sup 18/O incorporation into the substrate. No exchange was found suggesting that the catalytic step is not followed by a higher energy transition state and that it or a step before it appears to be rate-limiting.

• Research Organization: Univ. of California at San Diego, La Jolla
• OSTI ID: 5283397
• Report Number(s): CONF-8606151-
• Conference Information: Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Journal Volume: 45:6
• Country of Publication: United States
• Language: English

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59 BASIC BIOLOGICAL SCIENCES
62 RADIOLOGY AND NUCLEAR MEDICINE
ALCOHOLS
AMINES
AMMONIUM COMPOUNDS
BIOCHEMICAL REACTION KINETICS
BLOOD COAGULATION FACTORS
BODY
CARBON 13
CARBON ISOTOPES
CARBOXYLESTERASES
CHEMICAL REACTIONS
CHEMICAL SHIFT
CHOLINE
CLEAVAGE
COAGULANTS
CRYSTAL STRUCTURE
DECOMPOSITION
DIGESTIVE SYSTEM
DRUGS
ENDOCRINE GLANDS
ENZYMATIC HYDROLYSIS
ENZYMES
ESTERASES
ESTERS
EVEN-EVEN NUCLEI
EVEN-ODD NUCLEI
GLANDS
HEMATOLOGIC AGENTS
HYDROLASES
HYDROLYSIS
HYDROXY COMPOUNDS
ISOTOPES
KINETICS
LIGHT NUCLEI
LIPASE
LIPIDS
LIPOTROPIC FACTORS
LYSIS
MAGNETIC RESONANCE
MICROSTRUCTURE
NMR SPECTRA
NUCLEAR MAGNETIC RESONANCE
NUCLEI
ORGANIC COMPOUNDS
ORGANIC PHOSPHORUS COMPOUNDS
ORGANS
OXYGEN 16
OXYGEN 18
OXYGEN ISOTOPES
PANCREAS
PHOSPHOLIPIDS
QUATERNARY COMPOUNDS
REACTION KINETICS
RESONANCE
SOLVOLYSIS
SPECTRA
STABLE ISOTOPES
SUBSTRATES
VENOMS