Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “HMM”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 91 records · Page 5Linked to original sources
PubMed1988

[Effect of Ca2+ on the motility of myosin head in the F-actin-HMM complex].

Using polarization microfluorimetry, the effect of Ca2+ on the mode of interaction between heavy meromyosin labeled with 1.5-IAEDANS and F-actin was studied. The results obtained suggest that both the orientation and motility of myosin heads in the F-actin-heavy meromyosin complex depend on Ca2+ concentration. The experimental results are suggestive of the existence of a myosin-coupled regulatory system in skeletal muscles.

Actins↗
PubMed1980

Kinetic studies with synthetic myosin minifilaments show the equivalence of actomyosin and acto-HMM ATPases.

The kinetic properties of actomyosin have been examined using complexes of actin with the recently described (Reisler, E., Smith, C., and Seegan, G. (1980) J. Mol. Biol., in press) short, bipolar synthetic myosin filaments (minifilaments). It is shown, in contrast to previous observations with aggregated and insoluble myosin, that the kinetic behavior of actomyosin is similar to that of acto-heavy meromyosin. Owing to their size, solubility, and stability under conditions of the actin-activated ATPase measurements, the minifilaments provide a well defined experimental system. Thus, they constitute a convenient and appropriate material for studying actomyosin interactions.

Actins↗
PubMed1990

The rigor configuration of smooth muscle heavy meromyosin trapped by a zero-length cross-linker.

When chicken gizzard heavy meromyosin (HMM) in its rigor complex with actin was reacted with the zero-length cross-linker 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDC), HMM cross-linked with actin but also the two heads of the HMM molecule cross-linked to each other [Onishi, H., Maita, T., Matsuda, G., & Fujiwara, K. (1989) Biochemistry 28, 1898-1904, 1905-1912]. By ultracentrifugal fractionation of the EDC-treated acto-HMM in the presence of Mg-ATP, we obtained a preparation enriched for gizzard HMM with cross-linked heads. When HMM molecules in this preparation were rotary-shadowed and observed in an electron microscope, many head pairs were in contact with each other. The amount of HMM with cross-linked heads determined by electron microscopy was equal to that of the cross-linked NH2-terminal 24K tryptic fragments of HMM heavy chains determined by NaDodSO4 gel electrophoresis, indicating that this cross-linking is primarily responsible for the contact observed between two HMM heads. Most pairs of the contacted heads originated in the same HMM molecule, although a few pairs belonged to different HMM molecules. Cross-linking between the two heads of the same HMM molecule appeared to occur within the distal, more globular half of each head. However, the cross-linking sites were located at different positions within the globular portion. The actin-activated Mg-ATPase activity of the HMM sample treated with EDC in the presence of actin increased in a biphasic manner, depending on the concentration of F-actin, with two apparent association constants: 2.9 x 10(4) M-1 and one much less than 1 x 10(4) M-1. Since the apparent association constant obtained with the HMM control was similar to the latter value, the association constant for HMM molecules with cross-linked heads was identified to be the former value. The binding of HMM to actin was thus strengthened at least by a factor of 3 by the cross-linking between two HMM heads. These results suggest that HMM heads are trapped by treatment with EDC in the rigor complex configuration and that this configuration is retained even after the HMM has been released from actin. The EDC reactivity of rabbit skeletal muscle HMM, however, was different from that of chicken gizzard HMM. The treatment of acto-HMM complexes with EDC did not generate cross-linking between two skeletal muscle HMM heads.

Actins↗
PubMed1972

Heavy meromyosin: evidence for a refractory state unable to bind to actin in the presence of ATP.

The binding of actin to heavy meromyosin (HMM) in the presence of ATP was studied by analytical ultracentrifuge and ATPase studies. At 0 degrees C, at very low ionic strength, the double-reciprocal plot of HMM ATPase against actin concentration is linear. If one assumes that all of the HMM is bound to actin when the ATPase activity equals V(max), then, at an actin concentration where the actin-HMM ATPase is 85% of V(max), all but 15% of the HMM should be complexed with actin. However, when the binding of HMM to actin in the presence of ATP was measured with the analytical ultracentrifuge, more than 60% of the HMM was not bound to actin. From experiments with EDTA- and Ca-ATPases it seemed unlikely that the unbound HMM was denatured. It is thus possible that during the steady-state hydrolysis of ATP, HMM spends more than 50% of its cycle of interaction with actin and ATP in a "refractory state," unable to bind to actin, i.e., while an HMM molecule goes through one cycle of interaction with actin and ATP, an actin monomer could bind and release several HMM molecules so that the turnover rate per mole of added actin would be considerably greater than that per mole of added HMM. Comparison of the rate of ATPase activity at very high actin concentration with that at very high HMM concentration shows that this is indeed so. Therefore, both kinetic and ultracentrifuge studies suggest that the HMM exists in a refractory state during a large part of its cycle of interaction with actin and ATP.

Actins↗
PubMed1987

Conformational changes in myosin and heavy meromyosin from chicken gizzard associated with phosphorylation.

Heavy meromyosin (HMM) undergoes a conformational transition between a rapidly and a slowly sedimenting form, during which it sediments as a single peak in the ultracentrifuge with sedimentation coefficients between 7.5 and 9S. Changes in sedimentation velocity and ATPase activity produced by changes in ionic strength, phosphorylation of HMM or addition of MgATP are interpreted in terms of equilibria between the rapidly and slowly sedimenting forms, the observed values of activity and sedimentation velocity being determined by the fraction of HMM in each form. Phosphorylation of the 20 kDa light chain or raising the ionic strength decrease the sedimentation velocity, by decreasing the fraction of HMM in the rapidly sedimenting form, while addition of ATP increases sedimentation velocity upon forming a 9S HMM-ADP-Pi complex. Electron microscopic studies support this interpretation showing the presence of two distinct conformations of HMM--extended and flexed, which correspond to the 7.5S and 9S forms, respectively (Suzuki et al., 1985). In samples prepared at high ionic strengths, the heads extend away from the tail in a more or less random orientation, while at low ionic strength, the molecule is flexed at the head-tail junction assuming a more compact structure, that appears to account for its more rapid sedimentation rate. The degradation rates of the heavy chain and the 20 kDa light chain of HMM on digestion with papain indicate the presence of three forms of HMM differing in their susceptibility to papain. At 25 mM NaCl, HMM is rapidly digested in the absence of ATP, while addition of ATP decreases digestibility by a factor of ten, upon formation of a complex of HMM with the products of ATP hydrolysis. Above 0.4 M NaCl, HMM is degraded at an intermediate rate that is not affected by ATP. When the ionic strength is varied, the rate of disappearance of the heavy chain depends linearly on the sedimentation velocity in both the phosphorylated and dephosphorylated states, indicating that the rate of proteolysis is determined primarily by the fraction of HMM in the rapidly and slowly sedimenting forms. The same pattern is seen in the disappearance of the 20 kDa light chain of dephosphorylated HMM on cleavage at a site 4 kDa from the N-terminus, indicating that the cleavage of the light chain also depends on the fraction of HMM in the rapidly and slowly sedimenting forms.(ABSTRACT TRUNCATED AT 400 WORDS)

Adenosine Triphosphatases↗
PubMed1990

Structure and function of the cytoskeleton of a Dictyostelium myosin-defective mutant.

To study the role of conventional myosin in nonmuscle cells, we determined the cytoskeletal organization and physiological responses of a Dictyostelium myosin-defective mutant. Dictyostelium hmm cells were created by insertional mutagenesis of the myosin heavy chain gene (De Lozanne, A., and J. A. Spudich. 1987. Science (Wash. DC). 236: 1086-1091). Western blot analysis using different mAbs confirms that hmm cells express a truncated myosin fragment of 140 kD (HMM-140 protein) instead of the normal 243-kD myosin heavy chain (MHC). Spontaneous revertants appear at a frequency less than 4 x 10(-5), which synthesize normal myosin and are capable of forming thick filaments. In hmm cells, the HMM-140 protein is diffusely distributed in the cytoplasm, indicating that it cannot assemble into thick filaments. The actin distribution in these mutant cells appears similar to that of wild-type cells. However, there is a significant abnormality in the organization of cytoplasmic microtubules, which penetrate into lamellipodial regions. The microtubule networks consist of approximately 13 microtubules on average and their pattern is abnormal. Although hmm cells can form mitotic spindles, mitosis is not coordinated with normal furrow formation. The hmm cells are clearly defective in the contractile events that lead to normal cytokinesis. The retraction of different regions of the cell can result in the occasional pinching off of part of the cell. This process is not coupled with formation of mitotic spindles. There is no specific accumulation of HMM-140 in such constrictions, whereas 73% of such cells show actin concentrated in these regions. The mutant hmm cells are also deficient in capping of Con-A-bound surface receptors, but instead internalize this complex into the cytoplasm. The hmm cells display active phagocytosis of bacteria. Whereas actin is concentrated in the phagocytic cups, HMM-140 protein is not localized in these regions. cAMP, a chemoattractant that induces drastic rounding up and formation of surface blebs in wild type cells, does not induce rounding up in the hmm cells. A Triton-permeabilized cell model of the wild-type amebae contracts on reactivation with Mg-ATP, whereas a model of the hmm cell shows no detectable contraction. Our data demonstrate that the conventional myosin participates in the significant cortical motile activities of Dictyostelium cells, which include rounding up, constriction of cleavage furrows, capping surface receptors, and establishing cell polarity.

Animals↗
PubMed1977

Effects of ATP and ADP on hydrogen-deuterium exchange in heavy meromyosin. An examination of difference spectra.

The exchange reaction of peptide hydrogens with deuterium has been followed by measuring the decrease of the amide II band for heavy meromyosin (HMM). The difference spectra between HMM and HMM + ATP, between HMM and HMM + ADP, and between HMM + ATP and HMM + ADP have been examined as functions of time in order to detect small differences in the kinetic behavior of these different states of HMM. It has been found that, at 14 degrees C and 26 degrees C (pH 8.0), the exchange reaction is slightly slower for HMM + ATP than for HMM, and slightly slower for HMM + ADP than for HMM + ATP. This indicates that the secondary structure of HMM changes its flexibility during the ATP splitting cycle.

Adenosine Diphosphate↗
PubMed1983

Calcium-sensitive binding of heavy meromyosin to regulated actin requires light chain 2 and the head-tail junction.

Sedimentation in a preparative ultracentrifuge was used to determine the affinity of heavy meromyosin, HMM, for regulated actin, F-actin plus troponin-tropomyosin, in the presence of MgATP at pH 7.0, 20 degrees C, and mu = 18 mM. HMM was prepared from vertebrate striated muscle myosin by a mild chymotryptic digestion. This HMM contained 85-90% intact 19 000-dalton light chains, LC2. In the presence of calcium, 90% of the HMM bound to regulated actin with an association constant of (2-4) X 10(4) M-1. In the absence of calcium, while one-third of the HMM bound with an affinity similar to that observed in the presence of calcium, the rest bound much more weakly. It was not possible to accurately determine the association constant for this weakly binding HMM, but a 20-fold reduction in affinity is consistent with the binding data. The binding of single-headed heavy meromyosin to regulated actin was similarly sensitive to the calcium concentration. Since removal of calcium inhibits the regulated actin-activated ATPase of HMM greater than 20-fold, troponin-tropomyosin must be capable of inhibiting both the binding of HMM to regulated actin and a step which occurs after binding but prior to product release. Removal of LC2 increased the fraction of HMM with calcium-insensitive binding, and adding LC2 back to this depleted HMM restored most of the calcium sensitivity. Chymotryptic cleavage of LC2 to a 17 000-dalton fragment destroyed the calcium-sensitive binding of HMM to regulated actin. Phosphorylation of LC2, however, had no detectable effect on this binding. Thus, the calcium-sensitive binding of HMM to regulated actin requires that both the head-tail junction and the N-terminal part of LC2 be intact. Binding studies with cross-linked regulated actins and kinetic measurements of the rates of change in turbidity demonstrate that this calcium sensitivity is due to calcium binding to troponin and not to LC2.

Actins↗