Roles of phosphorylation and nucleotide binding domains in calcium transport by sarcoplasmic reticulum adenosinetriphosphatase

Journal Article · Tue Aug 09 04:00:00 EDT 1988 · Biochemistry; (United States)

OSTI ID:6829461

Teruel, J A; Inesi, G

The roles of the phosphorylation (phosphorylated enzyme intermediate) and nucleotide binding domains in calcium transport were studied by comparing acetyl phosphate and ATP as substrates for the Ca/sup 2 +/-ATPase of sarcoplasmic reticulum vesicles. The authors found that the maximal level of phosphoenzyme obtained with either substrate is approximately 4 nmol/mg of protein, corresponding to the stoichiometry of catalytic sites in their preparation. The initial burst of phosphoenzyme formation observed in the transient state, following addition of either substrate, is accompanied by internalization of 2 mol of calcium per mole of phosphoenzyme. The internalized calcium is then translocated with a sequential pattern, independent of the substrate used. Following a rate-limiting step, the phosphoenzyme undergoes hydrolytic cleavage and proceeds to the steady-state activity which is soon back inhibited by the rise of Ca/sup 2 +/ concentration in the lumen of the vesicles. When the back inhibition is released by the addition of oxalate, substrate utilization and calcium transport occur with a ratio of 1:2, independent of the substrate and its concentration. When the nucleotide binding site is derivatized with FITP, the enzyme can still utilize acetyl phosphate (but not ATP) for calcium transport. These observations demonstrate that the basic coupling mechanism of catalysis and calcium transport involves the phosphorylation and calcium binding domains, and not the nucleotide binding domain. On the other hand, occupancy of the FITC-sensitive nucleotide site is involved in kinetic regulation not only with respect to utilization of substrate for the phosphoryl transfer reaction but also for subsequent steps related to calcium translocation and phosphoenzyme turnover.

• Research Organization: Univ. of Maryland School of Medicine, Baltimore (USA)
• OSTI ID: 6829461
• Journal Information: Biochemistry; (United States), Journal Name: Biochemistry; (United States) Vol. 27:16; 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
ATP-ASE
BETA DECAY RADIOISOTOPES
BETA-MINUS DECAY RADIOISOTOPES
BIOCHEMICAL REACTION KINETICS
CALCIUM 45
CALCIUM CHLORIDES
CALCIUM COMPOUNDS
CALCIUM HALIDES
CALCIUM ISOTOPES
CATIONS
CELL CONSTITUENTS
CHARGED PARTICLES
CHEMICAL REACTIONS
CHLORIDES
CHLORINE COMPOUNDS
DAYS LIVING RADIOISOTOPES
ENZYME ACTIVITY
ENZYMES
EVEN-ODD NUCLEI
HALIDES
HALOGEN COMPOUNDS
HYDROLASES
INTERMEDIATE MASS NUCLEI
IONS
ISOTOPES
KINETICS
LIGHT NUCLEI
MEMBRANE TRANSPORT
NUCLEI
NUCLEOTIDES
ODD-ODD NUCLEI
ORGANIC COMPOUNDS
ORGANOIDS
PHOSPHOHYDROLASES
PHOSPHORUS 32
PHOSPHORUS ISOTOPES
PHOSPHORYLATION
RADIOISOTOPES
REACTION KINETICS
SARCOPLASMIC RETICULUM