Ca<sup>2+</sup>-Induced Conformational Change of Troponin C from the Japanese Pearl Oyster, <i>Pinctada fucata</i>

Abstract

Troponin is a thin filament-associated regulator of vertebrate striated muscle contraction. Troponin changes its structure upon Ca 2+ binding to troponin C, one of the subunits of troponin, allowing myosin to interact with actin. We recently elucidated the molecular characteristics of the Japanese pearl oyster Pinctada fucata troponin C (Pifuc-TnC), revealing the possibilities that Pifuc-TnC and vertebrate muscle TnC play dissimilar roles in muscle contraction. Pifuc-TnC has four EF-hand motifs, but, unlike vertebrate TnC, only one (site IV) was predicted to bind Ca 2+. To confirm the number of Ca 2+-binding sites in Pifuc-TnC and whether Ca 2+ binding induces a conformational change, we purified the full-length protein and a variant, Pifuc-TnC-E142Q (that has a mutation in the predicted Ca 2+-binding site of site IV), following their expression in laboratory E. coli. Isothermal titration calo

Tropomyosin Isoform Expression in the Adductor Muscle of the Japanese Pearl Oyster, <i>Pinctada fucata</i>

“…We recently showed that TnC, a subunit of Tn, might participate in the regulation of phasic muscle by virtue of its molecular characterization [5] [19]. In this study, DSC analysis clearly revealed that the types of TM isolated from phasic and catch muscles are different ( Figure 6), raising the possibility that Tn might interact with Pifuc-TM-1.…”

“…It was reported in a scallop striated muscle study that the movement of TM was induced by the Ca 2+ concentration, as it is in vertebrates [6]. Pifuc-TM-2 might be involved in phasic muscle regulation in collaboration with Tn, which is expressed mainly in phasic muscle [5] [19].…”

Molecular Cloning and Tissue Distribution of Troponin I from the Japanese Pearl Oyster, <i>Pinctada fucata</i>

“…Our studies have focused on the adductor muscle Tn subunits to investigate the molecular mechanism of thin filament-linked regulation of molluscan muscle contraction. To this end, we recently characterized P. fucata TnC (Pifuc-TnC) [12] [13]. Pifuc-TnC has four EF-hand motifs, sites I-IV, and, like other molluscan TnCs [14] [15] [16], only site IV is able to bind Ca 2+ [12].…”

“…To this end, we recently characterized P. fucata TnC (Pifuc-TnC) [12] [13]. Pifuc-TnC has four EF-hand motifs, sites I-IV, and, like other molluscan TnCs [14] [15] [16], only site IV is able to bind Ca 2+ [12]. A conformational change is induced in Pifuc-TnC upon Ca 2+ -binding [12].…”

“…Pifuc-TnC has four EF-hand motifs, sites I-IV, and, like other molluscan TnCs [14] [15] [16], only site IV is able to bind Ca 2+ [12]. A conformational change is induced in Pifuc-TnC upon Ca 2+ -binding [12]. A predicted three-dimensional model of Pifuc-TnC closely resembled that of vertebrate TnC, apart from a short loop (four amino acids) in the former structure that breaks the α-helix connecting the Nand C-terminal lobes [13].…”