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Rabbit M type phosphoglyceromutase: comparative effects of two thiol reagents antibody reaction and hybridization
M O Prehu1, C Prehu, M C Calvin
1Inserm U.91, Hôpital Henri Mondor, Crétil, France.
Abstract:
1. Treatment of purified rabbit phosphoglyceromutase (M type) with N-ethylmaleimide or with iodoacetamide produces the concurrent loss of phosphoglyceromutase activity with its collateral glycerate-2,3-P2 phosphatase activity. 2. Differences are observed in the protective effect of glycerate-2,3-P2 and of glycolate-2-P against N-ethylmaleimide and iodoacetamide treatments. 3. Specific chicken antibodies obtained by injection of the purified rabbit M type phosphoglyceromutase do not cross-react with the B type but neutralize both rabbit and human M type phosphoglyceromutase. 4. Purified rabbit M type phosphoglyceromutase can hybridize in vitro with the purified human B type or with purified human glycerate-2,3-P2 synthase. 5. Its ability to hybridize with glycerate-2,3-P2 synthase is unchanged after iodoacetamide treatment.
Insights
Treatment with N-ethylmaleimide or iodoacetamide inactivated rabbit phosphoglyceromutase (M type) and its phosphatase activity. Antibodies neutralized M type enzymes, while hybridization revealed distinct interactions with human enzymes.
Area of Science:
- Biochemistry
- Enzymology
Background:
- Phosphoglyceromutase (PGM) is crucial in glycolysis.
- M type PGM exhibits collateral glycerate-2,3-P2 phosphatase activity.
- Understanding PGM's structure-function relationship is vital.
Purpose of the Study:
- To investigate the enzymatic activities of rabbit M type phosphoglyceromutase.
- To explore the effects of chemical modification on enzyme activity.
- To characterize antibodies against M type PGM and its cross-reactivity.
Main Methods:
- Purification of rabbit M type phosphoglyceromutase.
- Enzyme activity assays using N-ethylmaleimide and iodoacetamide.
- Antibody production and cross-reactivity testing.
- In vitro hybridization assays with human enzymes.
Main Results:
- N-ethylmaleimide and iodoacetamide inactivated both phosphoglyceromutase and phosphatase activities.
- Glycerate-2,3-P2 and glycolate-2-P showed differential protection against chemical treatments.
- Chicken antibodies neutralized M type PGM but not B type.
- Rabbit M type PGM hybridized with human B type PGM and glycerate-2,3-P2 synthase, retaining this ability after iodoacetamide treatment.
Conclusions:
- Chemical modification affects both M type PGM and its phosphatase activity.
- M type PGM possesses distinct antigenic properties compared to B type.
- The catalytic site involved in hybridization with glycerate-2,3-P2 synthase is distinct from the iodoacetamide-reactive site.