Binding of Ca/sup 2 +/ to the calcium adenosinetriphosphatase of sarcoplasmic reticulum

Journal Article · Tue Nov 15 04:00:00 EST 1988 · Biochemistry; (United States)

OSTI ID:6245108

Petithory, J R; Jencks, W P

The binding of Ca/sup 2 +/ and the resulting change in catalytic specificity that allows phosphorylation of the calcium ATPase of sarcoplasmic reticulum by ATP were examined by measuring the amount of phosphoenzyme formation from (/sup 32/P)ATP, or /sup 45/Ca incorporation into vesicles, after the simultaneous addition of ATP and EGTA at different times after mixing enzyme and Ca/sup 2 +/. A burst of calcium binding in the presence of high (Ca/sup 2 +/) gives /approximately/ 12% phosphorylation and internalization of two Ca/sup 2 +/ at very short times after the addition of Ca/sup 2 +/ with this assay. This shows that calcium binding sites are available on the cytoplasmic-facing side of the free enzyme. Calcium binding to these sites induces the formation of /sup c/E/times/Ca/sub 2/, the stable high-affinity form of the enzyme, with k = 40 s/sup -1/ at saturating (Ca/sup 2 +/) and a half-maximal rate at approximately 20 ..mu..M Ca/sup 2 +/. The formation of /sup c/E/times/Ca/sub 2/ through a high-affinity pathway is described. The rate of exchange of /sup 45/Ca into the inner position of /sup c/E/times/Ca/sub 2/ shows and induction period and is not faster than the approach to equilibrium starting with E and /sup 45/Ca. The dissociation of /sup 45/Ca from the inner position of /sup c/E/times//sup 45/Ca/times/Ca in the presence of 3.2 ..mu..M Ca/sup 2 +/ occurs with a rate constant of 7 s/sup -1/. These results are inconsistent with a slow conformational change of free E to give /sup c/E, followed by rapid binding-dissociation of Ca/sup 2 +/.

• Research Organization: Brandeis Univ., Waltham, MA (USA)
• OSTI ID: 6245108
• Journal Information: Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 27:23; ISSN BICHA
• Country of Publication: United States
• Language: English

Related Subjects

550201* -- Biochemistry-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
ACID ANHYDRASES
ALKALINE EARTH ISOTOPES
ALKALINE EARTH METAL COMPOUNDS
ATP-ASE
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
CALCIUM 45
CALCIUM COMPOUNDS
CALCIUM ISOTOPES
CATIONS
CELL CONSTITUENTS
CHARGED PARTICLES
CHEMICAL REACTIONS
CONFORMATIONAL CHANGES
DAYS LIVING RADIOISOTOPES
ENDOPLASMIC RETICULUM
ENZYMES
EVEN-ODD NUCLEI
HYDROLASES
INTERMEDIATE MASS NUCLEI
IONS
ISOTOPES
KINETICS
LIGHT NUCLEI
NUCLEI
ODD-ODD NUCLEI
ORGANOIDS
PHOSPHOHYDROLASES
PHOSPHORUS 32
PHOSPHORUS ISOTOPES
PHOSPHORYLATION
RADIOISOTOPES
REACTION KINETICS
UPTAKE