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Kinetic evidence for two nucleotide binding sites on the CaATPase of sarcoplasmic reticulum

Journal Article · · Biochemistry; (United States)
DOI:https://doi.org/10.1021/bi00220a002· OSTI ID:5596297
;  [1]
  1. Univ. of the Pacific, San Francisco, CA (USA)

The CaATPase of sarcoplasmic reticulum was reacted with ({gamma}-{sup 32}P)ATP to form the covalent phosphoenzyme intermediate. Noncompetitive inhibition by reactive red-120 and chelation of calcium allowed the authors to monitor single-turnover kinetics of the phosphoenzyme reacting with water or added ADP at 0C. When ADP was added and the amount of product, ({gamma}-{sup 32}P)ATP, formed was measured, they found that added cold ATP did not interfere with the phosphoenzyme reacting with ADP. The authors conclude that ATP cannot bind where ADP binds, the phosphorylated active site. This implies that when ATP at high concentrations causes an acceleration of phosphoenzyme hydrolysis, it must do so by binding to an allosteric site. Considering the monoexponential nature of product formation they observed, simple one-nucleotide-site models cannot account for the above result.

OSTI ID:
5596297
Journal Information:
Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 30:6; ISSN 0006-2960; ISSN BICHA
Country of Publication:
United States
Language:
English