Phosphatidylinositol-specific phospholipase C from Bacillus cereus combines intrinsic phosphotransferase and cyclic phosphodiesterase activities: A sup 31 P NMR study
- Univ. of Oregon, Eugene (USA)
The inositol phosphate products formed during the cleavage of phosphatidylinositol by phosphatidylinositol-specific phospholipase C from Bacillus cereus were analyzed by {sup 31}P NMR. {sup 31}P NMR spectroscopy can distinguish between the inositol phosphate species and phosphatidylinositol. Chemical shift values (with reference to phosphoric acid) observed are {minus}0.41, 3.62, 4.45, and 16.30 ppm for phosphatidylinositol, myo-inositol 1-monophosphate, myo-inositol 2-monophosphate, and myo-inositol 1,2-cyclic monophosphate, respectively. It is shown that under a variety of experimental conditions this phospholipase C cleaves phosphatidylinositol via an intramolecular phosphotransfer reaction producing diacylglycerol and D-myo-inositol 1,2-cyclic monophosphate. The authors also report the new and unexpected observation that the phosphatidylinositol-specific phospholipase C from B. cereus is able to hydrolyze the inositol cyclic phosphate to form D-myo-inositol 1-monophosphate. The enzyme, therefore, possesses phosphotransferase and cyclic phosphodiesterase activities. The second reaction requires thousandfold higher enzyme concentrations to be observed by {sup 31}P NMR. This reaction was shown to be regiospecific in that only the 1-phosphate was produced and stereospecific in that only D-myo-inositol 1,2-cyclic monophosphate was hydrolyzed. Inhibition with a monoclonal antibody specific for the B.cereus phospholipase C showed that the cyclic phosphodiesterase activity is intrinsic to the bacterial enzyme. They propose a two-step mechanism for the phosphatidyl-inositol-specific phospholipase C from B. cereus involving sequential phosphotransferase and cyclic phosphodiesterase activities. This mechanism bears a resemblance to the well-known two-step mechanism of pancreatic ribonuclease, RNase A.
- OSTI ID:
- 5920643
- Journal Information:
- Biochemistry; (USA), Vol. 29:35; ISSN 0006-2960
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
PHOSPHODIESTERASES
NUCLEAR MAGNETIC RESONANCE
PHOSPHOTRANSFERASES
BACILLUS SUBTILIS
CHEMICAL SHIFT
INOSITOL
LIPASES
MONOCLONAL ANTIBODIES
NMR SPECTRA
PHOSPHORUS 31
ANTIBODIES
BACILLUS
BACTERIA
CARBOHYDRATES
CARBOXYLESTERASES
ENZYMES
ESTERASES
HYDROLASES
INOSITOLS
ISOTOPES
LIGHT NUCLEI
MAGNETIC RESONANCE
MICROORGANISMS
MONOSACCHARIDES
NUCLEI
ODD-EVEN NUCLEI
ORGANIC COMPOUNDS
PHOSPHORUS ISOTOPES
PHOSPHORUS-GROUP TRANSFERASES
RESONANCE
SACCHARIDES
SPECTRA
STABLE ISOTOPES
TRANSFERASES
550601* - Medicine- Unsealed Radionuclides in Diagnostics