Interactive properties of calmodulin

Cox, Jos A.

doi:10.1042/bj2490621

Cited by 114 publications

(47 citation statements)

References 71 publications

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“…Compared with the estimated dissociation constant of Ϸ1-5 M for the two higher affinity sites of unbound Ca 2ϩ -CaM (17,18), these results suggest that calcium binds at least 10-fold more weakly to bound CaM than to free CaM. By the linked equilibrium in Fig.…”

Section: Discussionmentioning

confidence: 72%

Regulation of Myosin V Processivity by Calcium at the Single Molecule Level

Lu¹,

Krementsova²,

Trybus³

2006

J. Biol. Chem.

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Calcium can affect myosin V (myoV) function in at least two ways. The full-length molecule, which adopts a folded inhibited conformation in EGTA, becomes extended and active in the presence of calcium. Calcium also dissociates one or more calmodulin molecules from the extended neck. Here we investigated at the single molecule level how calcium regulates the processive run length of full-length myosin V (dFull) and a truncated dimeric construct (dHMM), which cannot adopt the folded conformation. The processivity of dFull and dHMM is tightly controlled by the calcium and calmodulin concentration, with shorter runs occurring at higher calcium concentration. The data indicate that a calcium-dependent dissociation of calmodulin from the neck region of myoV terminates its processive run. dFull showed unexpected processive movement in EGTA, suggesting that a small population of extended, active molecules are in equilibrium with the inhibited, folded form. Single turnover assays showed that the ATPase activity of the folded full-length molecule is inhibited by more than 50-fold compared with the extended molecule. The results imply that activation and termination of the processive runs of myoV can be accomplished by multiple mechanisms. Myosin V (myoV)2 is a processive motor capable of taking multiple steps on actin filaments before dissociation (1). A striking feature of the molecule is its extended neck region which binds 6 calmodulins (CaM) and acts as a lever arm. This long neck region enables myoV to take 36-nm steps along the actin filament through a hand-over-hand mechanism (2), mediated by strain between the two heads (reviewed in Ref. 3). In addition to the widely accepted mechanical role of the lever arm, it is likely that the neck may also be involved in the regulation of myoV activity, given that CaM is a universal calcium sensor. Each CaM binds to an "IQ motif," with a consensus sequence of IQXXXRGXXXR, where X denotes any amino acid.In the absence of calcium, the IQ motifs are fully occupied by CaM. In the presence of calcium, one or more CaM molecules dissociate from the neck (4 -6). Based on data obtained with truncated neck constructs, it is likely that calcium-CaM can dissociate from the second IQ motif relative to the motor domain (5, 7).The actin-activated ATPase activity of full-length myoV, but not of shorter dimeric constructs, is inhibited in the absence of calcium and activated in its presence. Accompanying the activity change is a conformational change from a folded (inhibited) to an extended (active) conformation, which is seen only with the full-length molecule (5,8,9). Recent three-dimensional reconstructions of crystalline arrays of the folded form of myoV revealed structural details of the inhibited state (10). In this model, the molecule bends at the junction of the sixth IQ motif and the ␣-helical coiled-coil, so that the cargo-binding globular tail docks onto the two motor domains. The site of interaction is near loop 1 at the entrance to the nucleotide binding pocket, thus providing a ...

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Section: Discussionmentioning

confidence: 72%

Regulation of Myosin V Processivity by Calcium at the Single Molecule Level

Lu¹,

Krementsova²,

Trybus³

2006

J. Biol. Chem.

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“…According to one hypothesis, the four Ca2' -binding sites of calmodulin have very similar affinities with little or no cooperativity among the sites [369]; at pH 7.0 and ionic strength 150 mM, the Ca2+-saturation curve fits an ideal Langmuir isotherm with four independent sites of equal affinities ( K d 10 pM). Cox [370] has however demonstrated that the experimental data can also fit a second model 13711, according to which the N-and C-terminal Ca2 '-binding pairs differ markedly in their affinities for Ca2+. This latter model is supported by kinetic and conformational studies.…”

Section: The Cu2mentioning

confidence: 99%

Structural and functional aspects of calcium homeostasis in eukaryotic cells

Pietrobon

Virgilio

Pozzan

1990

European Journal of Biochemistry

203

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The maintenance of a low cytosolic free-Ca2+ concentration, ([Ca2+Ii) is a common feature of all eukaryotic cells. For this purpose a variety of mechanisms have developed during evolution to ensure the buffering of Ca2+ in the cytoplasm, its extrusion from the cell and/or its accumulation within organelles. Opening of plasma membrane channels or release of Ca2+ from intracellular pools leads to elevation of [Ca2+Ii; as a result, Ca2+ binds to cytosolic proteins which translate the changes in [Ca2+Ii into activation of a number of key cellular functions.The purpose of this review is to provide a comprehensive description of the structural and functional characteristics of the various components of [CaZ+li homeostasis in eukaryotes.It is well known how Sidney Ringer recognized the role of Ca2+ in muscle contraction [l] and how fortunate it was that he was working with frog heart muscle, rather than with skeletal muscle, given that contractility in the latter muscle is basically independent of extracellular Ca2 + . Less appreciated is the fact that it took a century before a wide-range test of the 'Ca2+ hypothesis' in the mediation of cellular responses became possible. So many years in fact elapsed between the publication of the seminal paper by Ringer in 1883 [l] and the report in 1982 by Tsien and co-workers [2] describing the application of quin 2 to the measurement of intracellular free Ca2+ ([Ca2++Ii) in small mammalian cells.The exploitation of fluorescent tetracarboxylate/pentacarboxylate dyes for measuring Ca2 + has enabled a thorough testing of Ca2+ involvement in a host of cellular responses spanning fertilization to muscle contraction, proliferation to neurotransmitter release and cell motility to cell death. The fluorescent dye technique has also complemented biochemistry, electrophysiology and molecular biology, and allowed the description in real time of the actual changes in [Ca2+Ii following the activation of membrane Ca2+ influx/ efflux pathways. Additional levels of complexity have emerged from the application of these techniques to the study of C a 2 + homeostasis and new problems have arisen, yet for the first Correspondewe to T. Pozzan, Institute of General Pathology, University of Ferrara, Via Borsari 46, 1-35121 Ferrara, ItalyAbbreviations. [Ca' +Ii, cytosolic free-Caz + concentration; VOC, voltage-operated channel; ROC, receptor-operated channel ; SMOC, second messenger-operated channcl; GOC, G-protein-operated channel; EC coupling, excitation-contraction coupling; MeDAsp, N-methyl u-aspartate; Ins(1 ,4,5)P3, inositol 1,4,5-trisphosphatc; Ins(1,3,4,5)P4, inositol 1,3,4,5-tetrakisphosphate; PtdIns(4,5)P2, phosphatidylinositol(4,5) bisphosphatc; SR, sarcoplasmic reticulum; ER. endoplasmic reticulum; TC, terminal cisternae; El. enzyme form 1 of ATPase; E2, enzyme form 2 of ATPase; G-protein, guanylnucleotide-binding protein.time we are beginning to understand CaZi signaling as the integration of multiple pathways.Ca" is maintained in the cytoplasm of mammalian cells at a concentration tha...

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“…3) [118, 1521. Ni2+ and Cd2+ selectively inhibit the Ca2+ response pointing to a calmodulin-mediated process since both cations are known to bind to calmodulin with high affinity [37]. Calmodulin is present in mammalian sperm [85] and sperm flagella [44, 861.…”

Section: Role Of Calciummentioning

confidence: 99%

Regulation of Mammalian Sperm Motility

Lindemann

Kanous

1989

Archives of Andrology

124

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The physiological regulation of sperm motility has become more amenable to investigation since the demonstration that CAMP and calcium play a role in modulating the functioning of the flagellar axoneme. Although the external triggering mechanisms that initiate motility and capacitation are still unknown, evidence supports a modification of the calcium balance by gated Ca2+ channels, accompanied by shifts in the internal pH. Ca2' and pH may in turn act indirectly through CAMP and CAMPdependent kinase (kinase,) to control the phosphorylation state of functional proteins in the flagellar axoneme. The role of calcium is of central importance, but it is clear that several separate Ca2+-dependent mechanisms are involved. Ca2+ controls the curvature of the sperm flagellum and, so, can change the motility of the sperm from progressive swimming to tumbling. Under the appropriate conditions, calcium appears to have the capacity to deactivate motility by activating phosphodiesterase and phosphatase. The deactivating effect of Ca2+ may be offset under some circumstances by coactivation of adenyl cyclase, so phosphorylation of the axoneme and the motility are maintained. The specific factors determining the predominant calcium effect are not yet known, but internal pH of the sperm may play a major role.Key Words: Sperm motility; Mammalian sperm; Mammalian reproduction; Flagellum; Calcium; pH. INTRODUCTIONThis review explores the influences regulating motility in mammalian sperm. Sperm motility depends on a flagellum, the organelle that develops the propulsive force for swimming. The cellular signals that modulate the action of the flagellar motor apparatus are the primary focus of this survey. Evidence is also examined to define the role of the male and female reproductive tracts in controlling sperm motility. To facilitate this review, structure/function relationships in mammalian sperm flagellum are briefly discussed. For the most part, the information included in this review has been obtained from research studies on mammalian sperm to identify the regulatory mechanisms most likely to be of physiological importance in mammalian reproduction. Received and accepted January 24, 1989. MORPHOLOGICAL PARAMETERSMammalian sperm swim by the propulsive action of a single flagellum. The structural foundation of the mammalian sperm flagellum is the 9 + 2 arrangement of microtubules that extends centrally through its entire length. This microtubule arrangement, called the axoneme, is common to all eukaryotic flagella ( Fig. 1) and also to cilia. Surrounding the axoneme are larger fibers, called coarse fibers or outer dense fibers (ODF), that are paired with the 9 outer doublet tubules. These fibers are anchored to the base of the flagellum in a caplike structure known as the connecting piece (CP). From there they taper in diameter along the flagellar length, terminating one-half to three-quarters of the way down the flagellum. The sperm mitochondria are wrapped around the basal one-third of the flagellum and this section is referred ...

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Interactive properties of calmodulin

Cited by 114 publications

References 71 publications

Regulation of Myosin V Processivity by Calcium at the Single Molecule Level

Regulation of Myosin V Processivity by Calcium at the Single Molecule Level

Structural and functional aspects of calcium homeostasis in eukaryotic cells

Regulation of Mammalian Sperm Motility

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