The Length of the A-M3 Linker Is a Crucial Determinant of the Rate of the Ca2+Transport Cycle of Sarcoplasmic Reticulum Ca2+-ATPase

Abstract This paper summarizes true equilibrium measurements for some partial reactions of the sarcoplasmic reticulum calcium pump transport cycle. The most important result is the estimation of the equilibrium constant for the interconversion of the two major conformational states of the protein, E (Ca2+ binding sites facing the cytoplasm) and E′ (Ca2+ binding sides facing the sarcoplasmic reticulum lumen). The value of K0 = [E′]/[E] cannot be evaluated directly by any method available at present, but observed cooperativity in the binding of Mg2+ and Ca2+ to unliganded protein strongly indictes that K0 ⪢ 1. The most probable value, valid within an order of magnitude, is K0 ≃ 103, i.e., the E′ state is more stable than the E state by about 4 kcal/mol.

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Equilibrium and Kinetic Studies of Calcium Transport and ATPase Activity in Sarcoplasmic Reticulum

Zeitschrift für Naturforschung C ◽

10.1515/znc-1982-7-819 ◽

1982 ◽

Vol 37 (7-8) ◽

pp. 685-691 ◽

Cited By ~ 30

Author(s):

G. Inesi ◽

M. Kurzmack ◽

D. Kosk-Kosicka ◽

D. Lewis ◽

H. Scofano ◽

...

Keyword(s):

Sarcoplasmic Reticulum ◽

Calcium Transport ◽

Inorganic Phosphate ◽

Calcium Release ◽

Kinetic Studies ◽

Hydrolytic Cleavage ◽

Kinetic Measurements ◽

Transport Cycle ◽

Rate Limiting ◽

Enzyme Intermediate

Abstract A number of equilibrium and kinetic measurements are presented to characterize the partial reactions of the ATPase and transport cycle in sarcoplasmic reticulum vesicles. The cycle begins with calcium and nucleotide binding on sites available on the outer surface of the vesicles. A phosphorylated enzyme intermediate is then formed, and the calcium sites are subjected to a change in their orientation and their affinity for calcium. It is shown that steps involved in calcium release on the inner side of the vesicles are rate limiting for the cycle, and are followed by hydrolytic cleavage of the intermediate with release of inorganic phosphate and recycling of the enzyme.

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Partial reactions in the catalytic and transport cycle of sarcoplasmic reticulum ATPase

Biochemistry ◽

10.1021/bi00616a023 ◽

1978 ◽

Vol 17 (23) ◽

pp. 5006-5013 ◽

Cited By ~ 114

Author(s):

S. Verjovski-Almeida ◽

M. Kurzmack ◽

G. Inesi

Keyword(s):

Sarcoplasmic Reticulum ◽

Transport Cycle

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Mechanism of Ca2+ transport by Ca2+-Mg2+-ATPase pump: analysis of major states and pathways

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1983.244.1.g3 ◽

1983 ◽

Vol 244 (1) ◽

pp. G3-G12 ◽

Cited By ~ 2

Author(s):

D. H. Haynes

Keyword(s):

Sarcoplasmic Reticulum ◽

Binding Site ◽

Random Order ◽

Product Inhibition ◽

Phosphate Binding ◽

Stoichiometric Amount ◽

Transport Cycle ◽

Adp Release ◽

Normal Transport ◽

A Charge

Mechanistic studies of Ca2+ transport by the Ca2+-Mg2+-ATPase of skeletal sarcoplasmic reticulum are reviewed, and a unifying model is proposed. The significant steps in the transport cycle are modeled in terms of occupation and disposition of three binding sites on the enzyme: a) two translocation sites capable of binding to Ca2+ or a charge-stoichiometric amount of alkali cation (M+) or H+, b) an ATP-ADP-binding site, and c) a phosphorylation or phosphate-binding site. The normal transport cycle is characterized as the following sequence of steps: a) binding of two Ca2+ and Mg-ATP to external sites with high affinity and random order, b) enzyme phosphorylation, c) inward translocation of the Ca2+-laden sites, d) Ca2+ release to the sarcoplasmic reticulum lumen and ADP release to the external medium (random order), e) binding of Mg2+ or a charge-stoichiometric amount of K+ plus H+ to the translocators, f) dephosphorylation, g) the return of the K+- and H+-laden translocators to the outside, and h) dissociation of K+ and H+ from the translocator and completion of the cycle with step a. The enzyme is characterized as a Ca2+-K+ plus H+ countertransporter. The K+ plus H+ remove Ca2+ from the inwardly oriented translocator, thereby relieving a product inhibition and increasing the rate of enzyme dephosphorylation.

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NMR relaxation measurements detect four intermediate states of ATPase and transport cycle of sarcoplasmic reticulum Ca2+-ATPase

Biochemical and Biophysical Research Communications ◽

10.1016/s0006-291x(88)81074-x ◽

1988 ◽

Vol 155 (1) ◽

pp. 236-242 ◽

Cited By ~ 7

Author(s):

Mary R. Klemens ◽

Charles M. Grisham

Keyword(s):

Sarcoplasmic Reticulum ◽

Nmr Relaxation ◽

Relaxation Measurements ◽

Intermediate States ◽

Transport Cycle

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A Reproducible Selective Stain for Cardiac Sarcoplasmic Reticulum

Proceedings, annual meeting, Electron Microscopy Society of America ◽

10.1017/s0424820100051086 ◽

1974 ◽

Vol 32 ◽

pp. 214-215

Author(s):

R. A. Waugh ◽

J. R. Sommer

Keyword(s):

Complex System ◽

Electron Microscope ◽

Sarcoplasmic Reticulum ◽

Transmission Electron Microscope ◽

Electron Microscopic ◽

Cardiac Sarcoplasmic Reticulum ◽

Transmission Electron

Cardiac sarcoplasmic reticulum (SR) is a complex system of intracellular tubules that, due to their small size and juxtaposition to such electron-dense structures as mitochondria and myofibrils, are often inconspicuous in conventionally prepared electron microscopic material. This study reports a method with which the SR is selectively “stained” which facilitates visualizationwith the transmission electron microscope.

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