Purification and characterization of the converting factor from E. coli chlA1
- Duke Univ., Durham, NC (United States)
Molybdopterin (MPT), a unique sulfur containing pterin which binds molybdenum, is a component of the molybdenum cofactor in all molybdenum enzymes except nitrogenase. In E. coli MPT is produced from a cyclic monophosphate precursor that lacks sulfur in a stoichiometric reaction with the converting factor. The converting factor activity has been ascribed to two proteins encoded at the chlA locus based on the ability of chlA mutants which lack converting factor activity to complement one another in vitro. The DNA sequence of the chlA locus indicates five open reading frames. The chlA1 mutant which is affected at gene 1 produces active converting factor. Purification of the converting factor through heat treatment, ammonium sulfate precipitation, and chromatography on DEAE cellulose, hydroxyapatite, and Mono Q resins yielded two proteins whose amino terminal amino acid sequences corresponded to the DNA sequences of genes 4 and 5 of the chlA locus. Although neither protein contains cysteine according to the DNA sequence, treatment of the chlA1 converting factor with iodoacetamide abolished converting factor activity. The activity could be restored by the addition of the purified small subunit of converting factor. These results suggest the gene 4 product contains a reactive sulfur necessary for converting factor activity.
- OSTI ID:
- 5307682
- Report Number(s):
- CONF-9104107-; CODEN: FAJOE
- Journal Information:
- FASEB Journal (Federation of American Societies for Experimental Biology); (United States), Vol. 5:4; Conference: 75. annual meeting of the Federation of American Societies for Experimental Biology (FASEB), Atlanta, GA (United States), 21-25 Apr 1991; ISSN 0892-6638
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
METALLOPROTEINS
CHEMICAL COMPOSITION
MOLYBDENUM
METABOLISM
AMINO ACID SEQUENCE
CHROMATOGRAPHY
FRACTIONATION
GENES
MUTANTS
ELEMENTS
METALS
MOLECULAR STRUCTURE
ORGANIC COMPOUNDS
PROTEINS
SEPARATION PROCESSES
TRANSITION ELEMENTS
560300* - Chemicals Metabolism & Toxicology