Monoclonal antibodies as probes of the topological arrangement of the two alpha subunits of the E coli RNA polymerase
Two monoclonal antibodies (mAb 124D1 and mAb 129C4) raised against the purified alpha subunit were noninhibitory in reactions directed by lacP/sup +/. Reconstitution of dissociated core polymerase in the presence of mAb 129C4 resulted in only a 5% recovery of enzyme activity whereas mAb 124D1 was without demonstrable effect. To determine the accessibility of the epitopes for the null mAbs and one inhibitory anti-alpha mAb 126C6 each was labeled with /sup 125/I. Following binding of an unlabeled mAb the plates were incubated with RNA polymerase and finally with one of the /sup 125/I-mAbs. The data indicate that in this assay two epitopes are available to mAb 126C6 in the RNA polymerase core or holoenzyme. Treatment of immobilized core enzyme with 2M or 4M urea resulted in an approximately two fold increase in the binding of /sup 125/I-mAb 129C4 or /sup 125/I-mAb 124D1 but lowered binding of /sup 125/I-mAb 126C6. The data suggest that the topology of the two alpha subunits in the native polymerase is such that one of the epitopes for mab 129C4 is blocked.
- Research Organization:
- Hunter College of CUNY, New York, NY
- OSTI ID:
- 5126358
- Report Number(s):
- CONF-8606151-
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Vol. 45:6; Conference: 76. annual meeting of the Federation of American Society for Experimental Biology, Washington, DC, USA, 8 Jun 1986
- Country of Publication:
- United States
- Language:
- English
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MONOCLONAL ANTIBODIES
BIOCHEMICAL REACTION KINETICS
MOLECULAR STRUCTURE
RNA POLYMERASES
ENZYME ACTIVITY
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IODINE 125
TRACER TECHNIQUES
ANTIBODIES
BACTERIA
BETA DECAY RADIOISOTOPES
DAYS LIVING RADIOISOTOPES
ELECTRON CAPTURE RADIOISOTOPES
ENZYMES
INTERMEDIATE MASS NUCLEI
IODINE ISOTOPES
ISOTOPE APPLICATIONS
ISOTOPES
KINETICS
MICROORGANISMS
NUCLEI
NUCLEOTIDYLTRANSFERASES
ODD-EVEN NUCLEI
PHOSPHORUS-GROUP TRANSFERASES
POLYMERASES
RADIOISOTOPES
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550201* - Biochemistry- Tracer Techniques