scholarly journals Photoaffinity labelling of smooth-muscle myosin by methylanthraniloyl-8-azido-ATP

1993 ◽  
Vol 292 (2) ◽  
pp. 439-444 ◽  
Author(s):  
S Maruta ◽  
M Ikebe
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Binding Sites ◽  
Smooth Muscle Myosin ◽  
Stable Complex ◽  
Atp Binding ◽  
Skeletal Muscle Myosin ◽  
Atp Binding Site ◽  
Tryptic Fragment ◽  
Muscle Myosin

Methylanthraniloyl-8-azido-ATP (Mant-8-N3-ATP), which binds to the 20 kDa C-terminal tryptic fragment of skeletal-muscle myosin subfragment-1 [Maruta, Miyanishi and Matsuda (1989) Eur. J. Biochem. 184, 213-221], was synthesized and used as a probe of the conformational change of smooth-muscle myosin. Mant-8-N3-ATP, like ATP, induced the formation of the 10 S conformation at low ionic strength. In the presence of vanadate, smooth-muscle myosin formed a stable complex with Mant-8-N3-ADP, and this complex showed the 10 S→6 S transition of myosin. ATP-binding sites for 6 S (extended state) and 10 S (folded state) myosin were studied by photolabelling of myosin with Mant-8-N3-ADP. For both 6 S and 10 S myosin, Mant-8-N3-ATP was incorporated into the 29 kDa N-terminal tryptic fragment of myosin heavy chain. This is unlike the labelling of skeletal-muscle myosin, in which the 20 kDa C-terminal fragment is labelled. The labelling of 29 kDa fragment was diminished significantly by addition of ATP. These results suggest that the conformation of the ATP-binding site of smooth-muscle myosin is different from that of skeletal-muscle myosin. To examine further the possible differences in the labelling site between 6 S and 10 S myosin, the affinity-labelled 29 kDa fragment was subjected to complete proteolysis by lysylendo-peptidase. The fluorescent-labelled-peptide map suggested that the Mant-8-N3-ADP-binding sites for 6 S and 10 S myosin were identical.

scholarly journals Assembly of smooth muscle myosin minifilaments: effects of phosphorylation and nucleotide binding.

1987 ◽  
Vol 105 (6) ◽  
pp. 3007-3019 ◽  
Author(s):  
K M Trybus ◽  
S Lowey
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Conformational Changes ◽  
Myosin Head ◽  
Smooth Muscle Myosin ◽  
Detectable Effect ◽  
Skeletal Muscle Myosin ◽  
Polar Type ◽  
Low Ionic Strength ◽  
Muscle Myosin

Small bipolar filaments, or "minifilaments," are formed when smooth muscle myosin is dialyzed against low ionic strength pyrophosphate or citrate/Tris buffers. Unlike synthetic filaments formed at approximately physiological ionic conditions, minifilaments are homogeneous as indicated by their hypersharp boundary during sedimentation velocity. Electron microscopy and hydrodynamic techniques were used to show that 20-22S smooth muscle myosin minifilaments are 380 nm long and composed of 12-14 molecules. By varying solvents, a continuum of different size polymers in the range of 15-30S could be obtained. Skeletal muscle myosin, in contrast, preferentially forms a stable 32S minifilament (Reisler, E., P. Cheung, and N. Borochov. 1986. Biophys. J. 49:335-342), suggesting underlying differences in the assembly properties of the two myosins. Addition of salt to the smooth muscle myosin minifilaments caused unidirectional growth into a longer "side-polar" type of filament, whereas bipolar filaments were consistently formed by skeletal muscle myosin. As with synthetic filaments, addition of 1 mM MgATP caused dephosphorylated minifilaments to dissociate to a mixture of folded monomers and dimers. Phosphorylation of the regulatory light chain prevented disassembly by nucleotide, even though it had no detectable effect on the structure of the minifilament. These results suggest that differences in filament stability as a result of phosphorylation are due largely to conformational changes occurring in the myosin head, and are not due to differences in filament packing.

Formation and properties of smooth muscle myosin 20-kDa light chain-skeletal muscle myosin hybrids and photocrosslinking from the maleimidylbenzophenone-labeled light chain to the heavy chain

1991 ◽  
Vol 288 (2) ◽  
pp. 584-590 ◽  
Author(s):  
Kallikat N. Rajasekharan ◽  
Jun-Ichiro Morita ◽  
Madhavan Mayadevi ◽  
Mitsuo Ikebe ◽  
Morris Burke
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Heavy Chain ◽  
Light Chain ◽  
Smooth Muscle Myosin ◽  
Skeletal Muscle Myosin ◽  
Muscle Myosin

scholarly journals Localization of the ATP-binding site in the 23-kDa and 20-kDa regions of the heavy chain of the skeletal muscle myosin head

1989 ◽  
Vol 184 (1) ◽  
pp. 213-221 ◽  
Author(s):  
Shinsaku MARUTA ◽  
Takayuki MIYANISHI ◽  
Genji MATSUDA
Keyword(s):  
Skeletal Muscle ◽  
Binding Site ◽  
Heavy Chain ◽  
Myosin Head ◽  
Atp Binding ◽  
Skeletal Muscle Myosin ◽  
Atp Binding Site ◽  
Muscle Myosin

IV. The mechanism of contraction - Introductory remarks

1973 ◽  
Vol 265 (867) ◽  
pp. 167-167
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Atpase Activity ◽  
Soluble Proteins ◽  
Smooth Muscle Myosin ◽  
Trypsin Treatment ◽  
Skeletal Muscle Myosin ◽  
Muscle Myosin

Our programme this afternoon is in two parts. We first welcome Professor Hamoir and Dr Kendrick-Jones to describe the several ways in which smooth muscle myosin differs from skeletal muscle myosin. It was in this biochemical field that my own work, with Dr Jennifer Williams, lay some twelve years ago. We were impressed at that time by the very low ATPase activity of the uterus actomyosin, and by the fact that on trypsin treatment meromyosins were obtained in some ways similar to those of skeletal muscle. Speakers this afternoon will have far more to tell us of the nature, behaviour and structure of the myosins concerned. We were also interested in the properties and possible function of certain soluble proteins, including tropomyosin, which figure so largely in smooth muscle constitution. This subject also will come up today.

scholarly journals Affinity labelling of smooth-muscle myosin light-chain kinase with 5′-[p-(fluorosulphonyl)benzoyl]adenosine

1993 ◽  
Vol 296 (1) ◽  
pp. 53-58 ◽  
Author(s):  
H Komatsu ◽  
M Ikebe
Keyword(s):  
Smooth Muscle ◽  
Binding Site ◽  
Light Chain ◽  
Myosin Light Chain Kinase ◽  
Myosin Light Chain ◽  
Smooth Muscle Myosin ◽  
Atp Binding ◽  
Atp Binding Site ◽  
Calmodulin Binding ◽  
Muscle Myosin

5′-(p-(Fluorosulphonyl)[14C]benzoyl)adenosine (FSBA) was synthesized and used as a probe to study the ATP-binding site of smooth-muscle myosin light-chain kinase (MLCK). FSBA modified both free MLCK and calmodulin/MLCK complex, resulting in inactivation of the kinase activity. Nearly complete protection of the calmodulin/MLCK complex against FSBA modification was obtained by addition of excess ATP whereas MLCK activity alone was lost in a dose-dependent manner even in the presence of excess ATP. These results suggest that FSBA modified ATP-binding sites and ATP-independent sites, and the latter sites are protected by calmodulin binding. The results also suggest that the ATP-binding site is accessible to the nucleotide substrate regardless of calmodulin binding. The FSBA-labelled MLCK was completely proteolysed by alpha-chymotrypsin, and the 14C-labelled peptides were isolated and sequenced. The sequence of the labelled peptide was Ala-Gly-X-Phe, where X is the labelled residue. The sequence was compared with the known MLCK sequence, and the labelled residue was identified as lysine-548, which is located downstream of the GXGXXG motif conserved among ATP-utilizing enzymes.

scholarly journals A comparison of the effects of calponin on smooth and skeletal muscle actomyosin systems in the presence and absence of caldesmon

1992 ◽  
Vol 288 (3) ◽  
pp. 733-739 ◽  
Author(s):  
S J Winder ◽  
C Sutherland ◽  
M P Walsh
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Smooth Muscle Actin ◽  
Smooth Muscle Myosin ◽  
Muscle Actin ◽  
Heavy Meromyosin ◽  
Muscle System ◽  
Skeletal Muscle Myosin ◽  
Actomyosin Atpase ◽  
Muscle Myosin

Thiosphosphorylated smooth muscle myosin and skeletal muscle myosin, both of which express Ca(2+)-independent actin-activated MgATPase activity, were used to examine the functional effects of calponin and caldesmon separately and together. Separately, calponin and caldesmon inhibited the actin-activated MgATPase activities of thiophosphorylated smooth muscle myosin and skeletal muscle myosin, calponin being significantly more potent in both systems. Calponin-mediated inhibition resulted from the interaction of calponin with actin since it could be reversed by increasing the actin concentration. Caldesmon had no significant influence on the calponin-induced inhibition of the smooth muscle actomyosin ATPase, nor did calponin have a significant effect on caldesmon-induced inhibition. In the skeletal muscle system, however, caldesmon was found to override the inhibitory effect of calponin. This difference probably reflects the lower affinity of skeletal muscle actin for calponin compared with that of smooth muscle actin. Calponin inhibition of skeletal muscle actin-activated myosin MgATPase was not significantly affected by troponin/tropomyosin, suggesting that the thin filament can readily accommodate calponin in addition to the troponin complex, or that calponin may be able to displace troponin. Calponin also inhibited acto-phosphorylated smooth muscle heavy meromyosin and acto-skeletal muscle heavy meromyosin MgATPases. The most appropriate protein preparations for analysis of the regulatory effects of calponin in the actomyosin system therefore would be smooth muscle actin, tropomyosin and thiophosphorylated myosin, and for analysis of the kinetic effects of calponin on the actomyosin ATPase cycle they would be smooth muscle actin, tropomyosin and phosphorylated heavy meromyosin, due to the latter's solubility.

scholarly journals Electron microscopy of synthetic myosin filaments. Evidence for cross-bridge. Flexibility and copolymer formation.

1975 ◽  
Vol 67 (1) ◽  
pp. 93-104 ◽  
Author(s):  
T D Pollard
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Structural Features ◽  
Length Frequency ◽  
Single Population ◽  
Skeletal Muscle Myosin ◽  
Distinctive Features ◽  
Myosin Filaments ◽  
The Mean ◽  
Muscle Myosin

Electron micrographs of negatively stained synthetic myosin filaments reveal that surface projections, believed to be the heads of the constituent myosin molecules, can exist in two configurations. Some filaments have the projections disposed close to the filament backbone. Other filaments have all of their projections widely spread, tethered to the backbone by slender threads. Filaments formed from the myosins of skeletal muscle, smooth muscle, and platelets each have distinctive features, particularly their lengths. Soluble mixtures of skeletal muscle myosin with either smooth muscle myosin or platelet myosin were dialyzed against 0.1 M KC1 at pH 7 to determine whether the simultaneous presence of two types of myosin would influence the properties of the filaments formed. In every case, a single population of filaments formed from the mixtures. The resulting filaments are thought to be copolymers of the two types of myosin, for several reasons: (a) their length-frequency distribution is unimodal and differs from that predicted for a simple mixture of two types of myosin filaments; (b) their mean length is intermediate between the mean lengths of the filaments formed separately from the two myosins in the mixture; (c) each of the filaments has structural features characteristic of both of the myosins in the mixture; and (d) their size and shape are determined by the proportion of the two myosins in the mixture.

Activation of smooth muscle myosin by smooth and skeletal muscle actins

FEBS Letters ◽  
1981 ◽  
Vol 134 (2) ◽  
pp. 197-202 ◽  
Author(s):  
Hanna Strzelecka-Golaszewska ◽  
Apolinary Sobieszek
Keyword(s):  
Skeletal Muscle ◽  
Smooth Muscle ◽  
Smooth Muscle Myosin ◽  
Muscle Myosin

scholarly journals Properties of the non-specific calcium-binding sites of rabbit skeletal-muscle myosin

1980 ◽  
Vol 185 (1) ◽  
pp. 265-268 ◽  
Author(s):  
J Wikman-Coffelt
Keyword(s):  
Skeletal Muscle ◽  
Light Chain ◽  
Binding Sites ◽  
Calcium Binding ◽  
Light Chains ◽  
Binding Properties ◽  
Skeletal Muscle Myosin ◽  
Heavy Chains ◽  
Calcium Binding Sites ◽  
Muscle Myosin

The non-specific Ca2+-binding sites of skeletal-muscle myosin are located on the light chains; with the dissociation of light chains there is a corresponding decrease in the number of Ca2+-binding sites on light-chain-deficient myosin. The released light chains have a decreased binding affinity. Myosin heavy chains indirectly influence the Ca2+-binding properties of light chains by increasing the affinity of light chains for bivalent cations; this influence varies with pH. Because of light-chain dissociation at low Ca2+ and/or Mg2+ concentrations, anomalies may exist when analyses of non-specific Ca2+-binding properties of myosin are assessed by dialysis equilibrium.

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