| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
J Biol Chem, Vol. 274, Issue 36, 25227-25236, September 3, 1999
From the Department of Chemical Pathology, University of Cape Town Medical School, 7925 Cape Town, South Africa
From the Banting and Best Department of Medical Research, C. H. Best Institute, University of Toronto, Toronto, Ontario M5G 1L6, Canada
From the Department of Physiology, University of Aarhus,
DK-8000 Aarhus C, Denmark
The nucleotide binding properties of mutants with
alterations to Asp351 and four of the other residues
in the conserved phosphorylation loop,
351DKTGTLT357, of sarcoplasmic reticulum
Ca2+-ATPase were investigated using an assay based on the
2',3'-O-(2,4,6-trinitrophenyl)-8-azidoadenosine triphosphate (TNP-8N3-ATP) photolabeling of
Lys492 and competition with ATP. In selected cases where
the competition assay showed extremely high affinity, ATP binding was
also measured by a direct filtration assay. At pH 8.5 in the absence of
Ca2+, mutations removing the negative charge of
Asp351 (D351N, D351A, and D351T) produced pumps that bound
MgTNP-8N3-ATP and MgATP with affinities 20-156-fold higher
than wild type (KD as low as 0.006 µM), whereas the affinity of mutant D351E was comparable
with wild type. Mutations K352R, K352Q, T355A, and T357A lowered the
affinity for MgATP and MgTNP-8N3-ATP 2-1000- and
1-6-fold, respectively, and mutation L356T completely prevented photolabeling of Lys492. In the absence of
Ca2+, mutants D351N and D351A exhibited the highest
nucleotide affinities in the presence of Mg2+ and at
alkaline pH (E1 state). The affinity of mutant D351A for MgATP was
extraordinarily high in the presence of Ca2+
(KD = 0.001 µM), suggesting a
transition state like configuration at the active site under these
conditions. The mutants with reduced ATP affinity, as well as mutants
D351N and D351A, exhibited reduced or zero CrATP-induced
Ca2+ occlusion due to defective CrATP binding.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  A. Marchand, A.-M. L. Winther, P. J. Holm, C. Olesen, C. Montigny, B. Arnou, P. Champeil, J. D. Clausen, B. Vilsen, J. P. Andersen, et al. Crystal Structure of D351A and P312A Mutant Forms of the Mammalian Sarcoplasmic Reticulum Ca2+-ATPase Reveals Key Events in Phosphorylation and Ca2+ Release J. Biol. Chem., May 23, 2008; 283(21): 14867 - 14882. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. D. Clausen, D. B. McIntosh, A. N. Anthonisen, D. G. Woolley, B. Vilsen, and J. P. Andersen ATP-binding Modes and Functionally Important Interdomain Bonds of Sarcoplasmic Reticulum Ca2+-ATPase Revealed by Mutation of Glycine 438, Glutamate 439, and Arginine 678 J. Biol. Chem., July 13, 2007; 282(28): 20686 - 20697. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. D. Clausen, D. B. McIntosh, D. G. Woolley, A. N. Anthonisen, B. Vilsen, and J. P. Andersen Asparagine 706 and Glutamate 183 at the Catalytic Site of Sarcoplasmic Reticulum Ca2+-ATPase Play Critical but Distinct Roles in E2 States J. Biol. Chem., April 7, 2006; 281(14): 9471 - 9481. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. B. McIntosh, J. D. Clausen, D. G. Woolley, D. H. MacLennan, B. Vilsen, and J. P. Andersen Roles of Conserved P Domain Residues and Mg2+ in ATP Binding in the Ground and Ca2+-activated States of Sarcoplasmic Reticulum Ca2+-ATPase J. Biol. Chem., July 30, 2004; 279(31): 32515 - 32523. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. Dode, J. P. Andersen, N. Leslie, J. Dhitavat, B. Vilsen, and A. Hovnanian Dissection of the Functional Differences between Sarco(endo)plasmic Reticulum Ca2+-ATPase (SERCA) 1 and 2 Isoforms and Characterization of Darier Disease (SERCA2) Mutants by Steady-state and Transient Kinetic Analyses J. Biol. Chem., November 28, 2003; 278(48): 47877 - 47889. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
D. Strugatsky, K.-E. Gottschalk, R. Goldshleger, E. Bibi, and S. J. D. Karlish Expression of Na+,K+-ATPase in Pichia pastoris: ANALYSIS OF WILD TYPE AND D369N MUTANT PROTEINS BY FE2+-CATALYZED OXIDATIVE CLEAVAGE AND MOLECULAR MODELING J. Biol. Chem., November 14, 2003; 278(46): 46064 - 46073. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
H. Ma, G. Inesi, and C. Toyoshima Substrate-induced Conformational Fit and Headpiece Closure in the Ca2+ATPase (SERCA) J. Biol. Chem., August 1, 2003; 278(31): 28938 - 28943. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. D. Clausen, D. B. McIntosh, B. Vilsen, D. G. Woolley, and J. P. Andersen Importance of Conserved N-domain Residues Thr441, Glu442, Lys515, Arg560, and Leu562 of Sarcoplasmic Reticulum Ca2+-ATPase for MgATP Binding and Subsequent Catalytic Steps: PLASTICITY OF THE NUCLEOTIDE-BINDING SITE J. Biol. Chem., May 23, 2003; 278(22): 20245 - 20258. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. B. McINTOSH, J. D. CLAUSEN, D. G. WOOLLEY, D. H. MacLENNAN, B. VILSEN, and J. P. ANDERSEN ATP Binding Residues of Sarcoplasmic Reticulum Ca2+-ATPase Ann. N.Y. Acad. Sci., April 1, 2003; 986(1): 101 - 105. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Liu and A. Barth Mapping Interactions between the Ca2+-ATPase and Its Substrate ATP with Infrared Spectroscopy J. Biol. Chem., March 14, 2003; 278(12): 10112 - 10118. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
L. R. Jones, R. L. Cornea, and Z. Chen Close Proximity between Residue 30 of Phospholamban and Cysteine 318 of the Cardiac Ca2+ Pump Revealed by Intermolecular Thiol Cross-linking J. Biol. Chem., July 26, 2002; 277(31): 28319 - 28329. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
J. D. Clausen, D. B. McIntosh, D. G. Woolley, and J. P. Andersen Importance of Thr-353 of the Conserved Phosphorylation Loop of the Sarcoplasmic Reticulum Ca2+-ATPase in MgATP Binding and Catalytic Activity J. Biol. Chem., September 14, 2001; 276(38): 35741 - 35750. [Abstract] [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| All ASBMB Journals | Molecular and Cellular Proteomics |
| Journal of Lipid Research | ASBMB Today |
