The Mechanics of Calcium Transport

Abstract

Abstract. With the recent atomic 1 models for the sarcoplasmic reticulum Ca 2+

The Molecular Basis for Cyclopiazonic Acid Inhibition of the Sarcoplasmic Reticulum Calcium Pump

“…In contrast, M4 is well defined, and its position is unchanged in comparison with previous structures (4,6). Electron cryomicroscopy of helical crystals revealed that TG binding causes a small downward movement of M3 and a restructuring of the M3/M4 loop (13,14). We believe that the position of M3 observed in previous structures (4,6) and in the helical crystals (13) is a characteristic of TG inhibition.…”

“…The potential drawbacks associated with these "static" images from crystallographic studies include structural changes caused by the inhibitors and nucleotide analogues used to stabilize the different conformational states of the enzyme. For instance, an electron cryomicroscopy structure originally visualized an exit pathway from the calcium transport sites to the lumen of the SR (11,12), and this channel was postulated to close in the presence of an inhibitor (13,14). However, a luminal exit channel has not been observed in the available crystal structures (4,6).…”

Characterizing Phospholamban to Sarco(endo)plasmic Reticulum Ca2+-ATPase 2a (SERCA2a) Protein Binding Interactions in Human Cardiac Sarcoplasmic Reticulum Vesicles Using Chemical Cross-linking

“…SERCA2a is a major Ca 2ϩ -handling protein in the heart that actively transports cytosolic Ca 2ϩ into the lumen of the SR, inducing myofilament relaxation (2). The dephosphorylated PLB monomer binds to and inhibits Ca 2ϩ -ATPase activity by decreasing the apparent Ca 2ϩ affinity of the enzyme (3,4), having little or no effect on the V max of the enzyme measured at saturating Ca 2ϩ concentration (1,5).…”

“…PLB monomers also oligomerize into noninhibitory homopentamers in the SR membrane (6) (stabilized by transmembrane Leu/Ile zipper interactions) (7), but the functional significance of the pentamer remains unclear. The inhibitory effect of PLB on SERCA2a activity is reversed when PLB is phosphorylated at Ser 16 (by protein kinase A) or Thr 17 (by calmodulin-dependent protein kinase) during ␤-adrenergic receptor stimulation of the heart, increasing the size of the contractile-dependent SR Ca 2ϩ -store and the rate of cardiac relaxation (1,2). Similarly, anti-PLB monoclonal antibodies binding near the phosphorylation site also reverse Ca 2ϩ pump inhibition by PLB (5,8), with functional effects indistinguishable from those of ␤-adrenergic stimulation (9).…”

“…Techniques assessing protein-protein interactions have included co-immunoprecipitation (11), chemical crosslinking (12,13,17), FRET (14,16,18), and two-dimensional * This work was supported, in whole or in part, by National Institutes of Health Grant HL49428. 1 To whom correspondence should be addressed: Krannert Institute of Cardiology, 1800 N. Capitol Ave., Indianapolis, IN 46202. co-crystallization (2,19), which have frequently given conflicting results. For example, some studies (including our own) suggest that the Ca 2ϩ pump with bound PLB is catalytically inactive, requiring PLB dissociation prior to enzyme activation (11,12,13,15,17), whereas others maintain that PLB is a subunit of the Ca 2ϩ pump that remains attached throughout the catalytic cycle (14,16,18).…”