Selective modification of phosphoribulokinase cysteines
The most reactive sulfhydryl in native phosphoribulokinase can be selectively alkylated with iodoacetate; complete loss of activity results. Composition and N-terminal analyses of the peptide bearing this -SH show that cys-16 is the modification site. In the presence of Mg and ATP, a nonessential -SH is modified; catalytic activity is unchanged. The peptide bearing this cysteine has been characterized by amino acid composition and N-terminal analyses (Phe-Phe-Asn-Pro-Val-Tyr-(Ile/Leu)...). Enzyme alkylated at this site is subject to reversible oxidative inactivation, showing that this cysteine is not involved in regulation of activity. A cysteine distinct from this nonessential residue is suggested by crosslinking studies to be close (5a) to cys-16. The site-directed inhibitor fluorosulfonylbenzoyladenosine (FSBA) inactivates enzyme by modifying cys-16. Carboxymethylation of enzyme after FSBA incorporation, followed by DTT treatment, unmasks the initial modification site and allows (/sup 14/C)-iodoacetate labeling. The tryptic peptide bearing this residue co-chromatographs with and is identical in composition to the cys-16 containing peptide.
- Research Organization:
- Medical College of Wisconsin, Milwaukee
- OSTI ID:
- 6180371
- Report Number(s):
- CONF-870644-
- Journal Information:
- Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States), Journal Name: Fed. Proc., Fed. Am. Soc. Exp. Biol.; (United States) Vol. 46:6; ISSN FEPRA
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
59 BASIC BIOLOGICAL SCIENCES
AMINO ACID SEQUENCE
AMINO ACIDS
BIOLOGICAL FUNCTIONS
CARBON 14 COMPOUNDS
CARBOXYLIC ACIDS
CHEMICAL COMPOSITION
CHEMICAL REACTIONS
CROSS-LINKING
CYSTEINE
ENZYME ACTIVITY
ENZYMES
FUNCTIONS
ISOTOPE APPLICATIONS
LABELLED COMPOUNDS
MOLECULAR STRUCTURE
ORGANIC ACIDS
ORGANIC COMPOUNDS
ORGANIC SULFUR COMPOUNDS
PEPTIDES
PHOSPHORUS-GROUP TRANSFERASES
PHOSPHOTRANSFERASES
POLYMERIZATION
PROTEINS
THIOLS
TRACER TECHNIQUES
TRANSFERASES