首页 | 本学科首页   官方微博 | 高级检索  
相似文献
 共查询到20条相似文献,搜索用时 15 毫秒
1.
Heat-induced gelation of myosin from leg and breast muscles of chicken was studied in 0.6 m KC1. Gel strength of breast myosin was higher than that of leg myosin between pH 5.2 and 6.0. Turbidity of breast myosin increased below pH 6.0 but that of leg myosin did not increase at pH 5.7. Turbidity of leg myosin was higher than that of breast myosin below pH 5.6. Viscosity of breast myosin increased between pH 5.5 and 6.0 as the pH decreases, although that of leg myosin decreased. The breast myosin assembled to form long filaments at pH 5.7, but leg myosin failed to form long filaments. At pH 5.4, breast myosin filaments became longer and leg myosin assembled into filaments though they were shorter than breast myosin filaments. The strength of heat-induced gel formed from the filamentous leg and breast myosins at acidic region was not influenced by F-actin. These results indicate that the strength of heat-induced gel of both myosins is closely related to their morphological properties.  相似文献   

2.
Regulation of embryonic smooth muscle myosin by protein kinase C   总被引:2,自引:0,他引:2  
Phosphorylation of the 20-kDa light chain regulates adult smooth muscle myosin; phosphorylation by the Ca2+/calmodulin-dependent enzyme myosin light chain kinase stimulates the actomyosin ATPase activity of adult smooth muscle myosin; the simultaneous phosphorylation of a separate site on the 20-kDa light chain by the Ca2+/phospholipid-dependent enzyme protein kinase C attenuates the myosin light chain kinase-induced increase in the actomyosin ATPase activity of adult myosin. Fetal smooth muscle myosin, purified from 12-day-old fertilized chicken eggs, is structurally different from adult smooth muscle myosin. Nevertheless, phosphorylation of a single site on the 20-kDa light chain of fetal myosin by myosin light chain kinase results in stimulation of the actomyosin ATPase activity of this myosin. Protein kinase C, in contrast, phosphorylates three sites on the fetal myosin 20-kDa light chain including a serine or threonine residue on the same peptide phosphorylated by myosin light chain kinase. Interestingly, phosphorylation by protein kinase C stimulates the actomyosin ATPase activity of fetal myosin. Moreover, unlike adult myosin, there is no attenuation of the actomyosin ATPase activity when fetal myosin is simultaneously phosphorylated by myosin light chain kinase and protein kinase C. These data demonstrate, for the first time, the in vitro activation of a smooth muscle myosin by another enzyme besides myosin light chain kinase and raise the possibility of alternate pathways for regulating smooth muscle myosin in vivo.  相似文献   

3.
P D Wagner  N D Vu 《Biochemistry》1988,27(17):6236-6242
The effects of light chain phosphorylation on the actin-activated ATPase activity and filament assembly of calf thymus cytoplasmic myosin were examined under a variety of conditions. When unphosphorylated and phosphorylated thymus myosins were monomeric, their MgATPase activities were not activated or only very slightly activated by actin, but when they were filamentous, their MgATPase activities were stimulated by actin. The phosphorylated myosin remained filamentous at lower Mg2+ concentrations and higher KC1 concentrations than did the unphosphorylated myosin, and the myosin concentration required for filament assembly was lower for phosphorylated myosin than for unphosphorylated myosin. By varying the myosin concentration, it was possible to have under the same assay conditions mostly monomeric myosin or mostly filamentous myosin; under these conditions, the actin-activated ATPase activities of the filamentous myosins were much greater than those of the monomeric myosins. The addition of phosphorylated myosin to unphosphorylated myosin promoted the assembly of unphosphorylated myosin into filaments. These results suggest that phosphorylation may regulate the actomyosin-based motile activities in vertebrate nonmuscle cells by regulating myosin filament assembly.  相似文献   

4.
Two types of myosin isolated from ordinary (fast) and dark (slow) muscles of carp were examined by ATPase and in vitro motility assays. Vmax of the ATPase activity and sliding velocity of ordinary myosin showed 1.6 and 1.5 times higher activities than those of dark myosin, whereas those of mammalian fast myosin were much higher, 3 to 10 times, than those of slow myosin. Although ordinary myosin had almost identical activities to those of mammalian fast myosin, activities of dark myosin was twice of those of mammalian slow myosin. This high motile activity of dark myosin can account for the physiological role of dark muscle in cruising of fish. By comparing Km of the actin-activated ATPase activity, ordinary myosin was appeared to have higher affinity to F-actin than dark myosin, and this was confirmed by the binding assay of HMM or S-1 of carp myosin to F-actin. Investigation of myosin assembly by electron microscopy and the centrifugation assay revealed that ordinary myosin assembled much poorly than dark myosin or mammalian fast myosin. This phenomenon may reflect characteristic cellular function of fish skeletal muscle.  相似文献   

5.
The rate of phosphorylation and dephosphorylation of smooth muscle myosin by myosin light chain kinase and by two myosin light chain phosphatases (gizzard phosphatase IV and aorta phosphatase) are measured in various conditions; the relationship between the rate of phosphorylation and dephosphorylation of myosin and the myosin conformation is also studied. The rate of dephosphorylation of myosin was completely inhibited in the presence of 1 mM MgCl2 and ATP at low ionic strength where phosphorylated myosin forms a folded conformation. The inhibition was released when myosin formed either an extended monomer or filaments. The rate of phosphorylation of myosin was also affected by the conformation of myosin. The rate for a folded myosin was slower than those for an extended monomer and filamentous myosin. The phosphorylation and dephosphorylation of heavy meromyosin, subfragment-1, and the isolated 20,000-dalton light chain are not inhibited at low ionic strength, and the rate of phosphorylation and dephosphorylation was decreased with increasing ionic strength. KCl dependence of the rate of phosphorylation and dephosphorylation of myosin was normalized by using KCl dependence of subfragment-1, and it was found that the marked inhibition of the rate of phosphorylation and dephosphorylation of myosin is closely related to the change from an extended to a folded conformation of myosin.  相似文献   

6.
Calponin (CaP), a thin filament-associated protein, plays an important role in the regulation of smooth muscle contractility. It has been known that CaP inhibits the actin-activated myosin MgATPase activity via binding to F-actin, and stimulates myosin MgATPase activity via binding to myosin. Our recent study revealed a new phenomenon that trace amount of CaP (TAC) could influence the function of different states of myosin. Our data showed that in the absence of actin, CaP, even in the concentration of 0.0001 microM, significantly increased the precipitations of 1 microM unphosphorylated myosin, Ca(2+)-CaM dependently, and independently phosphorylated myosin by MLCK, and stimulated the MgATPase activities of these myosins slightly but significantly. However, no obvious change of precipitation of myosin phosphorylated by PKA was observed, indicating the relative selective effect of TAC. In the presence of actin, myosin, and TAC, the increase of myosin precipitation was abolished, and no obvious changes of actin precipitations and actin-activated myosin MgATPase activities were observed implicating the highly efficiency of TAC on myosin being present in the absence of actin. Although we cannot give conclusive comments to our results, we propose that the high efficiency of TAC-myosin interaction is present in the regulation of the function of myosin when actin is dissociated from myosin, even if CaP/myosin ratio is very low; this high efficient interaction between TAC and myosin can be abolished by actin. However, why and how TAC can possess such a high efficiency to influence myosin and how the physiological significance of the high efficiency of TAC is in regulating the interaction between myosin and actin remain to be investigated.  相似文献   

7.
The ATPase activity of myosin and contraction time in extensor digitorum longus muscle, soleus muscle and cardiac muscle was compared in mammals differing in size. It was shown that the myosin ATPase activity of homologous muscles decreases and contraction time increases with increasing size of animals. The rate of tryptic digestion of myosin, the electrophoretic pattern of light chains of myosin and the effect of p-chloromercuribenzoate on ATPase activity of myosin were also studied. All these three myosin properties are very characteristic when the myosin from a fast muscle is compared with the myosin from a slow muscle of the same animal, but no relationship between these three myosin properties and ATPase activity of myosin was found, when homologous muscles of various mammals were compared.  相似文献   

8.
It is known that melanophilin is a myosin Va-targeting molecule that links myosin Va and the cargo vesicles in cells. Here we found that melanophilin directly activates the actin-activated ATPase activity of myosin Va and thus its motor activity. The actin-activated ATPase activity of the melanocyte-type myosin Va having exon-F was significantly activated by melanophilin by 4-fold. Although Rab27a binds to myosin Va/melanophilin complex, it did not affect the melanophilin-induced activation of myosin Va. Deletion of the C-terminal actin binding domain and N-terminal Rab binding domain of melanophilin resulted in no change in the activation of the ATPase by melanophilin, indicating that the myosin Va binding domain (MBD) is sufficient for the activation of myosin Va. Among MBDs, the interaction of MBD-2 with exon-F of myosin Va is critical for the binding of myosin Va and melanophilin, whereas MBD-1 interacting with the globular tail of myosin Va plays a more significant role in the activation of myosin Va ATPase activity. This is the first demonstration that the binding of the cargo molecule directly activates myosin motor activity. The present finding raises the idea that myosin motors are switched upon their binding to the cargo molecules, thus avoiding the waste of ATP consumption.  相似文献   

9.
An antiserum specific to dog myocardial myosin has been developed against highly purified myosin heavy chains. The antiserum is specific for the heavy chains of myosin, giving a single precipitin line in an immunodiffusion assay for either the heavy chains of myosin or native myosin, and does not react with any other myocardial proteins. In such assays myosin acts as a single, uniform antigen. Using this antiserum, a radioimmunoassay has been developed to quantitate myosin in a homogenate of myocardial tissue containing free myosin dissociated from other cellular components.  相似文献   

10.
1. The Ca2(+)-activated and Mg2+ actin-activated myosin ATPase activities of flightless mfd- mutant Drosophila flight muscle myosin were one-half and one-third of those of the wild-type fly muscle myosin, respectively. 2. In the two-dimensional gel electrophoresis, the spots corresponding to phosphorylated myosin light chains, Lfl and Ltl, were hardly detected in mfd- mutant myosin. 3. These results support not only the conclusion that phosphorylation of myosin light chains regulates Drosophila myosin ATPase activity but also the assumption that the phosphorylation of myosin light chains is directly involved in flight function of the Drosophila fly.  相似文献   

11.
在有Ca2+和钙调蛋白存在时,肌球蛋白轻链激酶催化肌球蛋白磷酸化,促使肌动蛋白激活的肌球蛋白(肌动球蛋白)Mg2+-ATP酶活性显著增加.然而,肌球蛋白磷酸化水平与Mg2+-ATP酶之间的关系是非线性的,原肌球蛋白可以进一步增加Mg2+-ATP酶的活性,但仍不改变它们之间的非线性关系.肌球蛋白轻链激酶的合成肽抑制剂抑制了肌球蛋白磷酸化和Mg2+-ATP酶活性,并导致平滑肌去膜肌纤维的等长收缩张力与速度的降低.结果提示肌球蛋白轻链激酶参与脊椎动物平滑肌收缩的调节过程,肌球蛋白轻链磷酸化作用会引起平滑肌收缩  相似文献   

12.
Myosin was purified from rabbit alveolar macrophages in a form that could not be activated by actin. This myosin could be phosphorylated by an endogenous myosin light chain kinase, up to 2 mol of phosphate being incorporated/mol of myosin. The site phosphorylated was located on the 20,000-dalton myosin light chain. Phosphorylation of macrophage myosin was found to be necessary for actin activation of myosin ATPase activity. Moreover, the actin-activated ATPase activity was found to vary directly with the extent of myosin phosphorylation, maximal phosphorylation (2 mol of Pi/mol of myosin) resulting in an actin-activated MgATPase activity of approximately 200 nmol of Pi/mg of myosin/min at 37 degrees C. These results establish that phosphyoyration of the 20,000-dalton light chain of myosin is sufficient to regulate the actin-activated ATPase activity of macrophage myosin.  相似文献   

13.
Changes in myosin isozymes during development of chicken breast muscle   总被引:1,自引:0,他引:1  
The patterns of myosin isozymes in embryonic and adult chicken pectoralis muscle were examined by electrophoresis in a non-denaturing gel system (pyrophosphate acrylamide gel electrophoresis), and both light chains and heavy chains of embryonic and adult myosin isozymes were compared. In pyrophosphate acrylamide gel electrophoresis, the predominant isozyme component in embryonic pectoralis myosin could be clearly distinguished from adult myosin isozymes. SDS-polyacrylamide gel electrophoresis indicated that the light chain composition of embryonic myosin was also different from that of adult myosin. The pattern of peptide fragments produced by myosin digestion with a-chymotrypsin differed significantly between embryonic and adult skeletal myosin. These results suggest that myosin in the embryonic pectoralis muscle is different in both light and heavy chain composition from myosin in the same adult tissue.  相似文献   

14.
Myosin was isolated from the ventricular myocardium of adult rats and the effect of time, 2-mercaptoethanol and inhibitors of proteases was investigated on its properties. It was found that the storage of cardiac muscle up to 4 hours does not influence the myosin ATPase, the electrophoretic pattern of light chains of myosin or the pattern of peptides produced by digestion of myosin with chymotrypsin. Neither does the presence of pepstatin and phenylmethyl sulfonylfluoride during myosin preparation influence the activity of myosin ATPase. It was found that the presence of 2-mercaptoethanol during myosin preparation enhances myosin ATPase of the product. This myosin was more stable when kept at 4 degrees C for four days.  相似文献   

15.
We have used electron microscopy and solubility measurements to investigate the assembly and structure of purified human platelet myosin and myosin rod into filaments. In buffers with ionic strengths of less than 0.3 M, platelet myosin forms filaments which are remarkable for their small size, being only 320 nm long and 10-11 nm wide in the center of the bare zone. The dimensions of these filaments are not affected greatly by variation of the pH between 7 and 8, variation of the ionic strength between 0.05 and 0.2 M, the presence or absence of 1 mM Mg++ or ATP, or variation of the myosin concentration between 0.05 and 0.7 mg/ml. In 1 mM Ca++ and at pH 6.5 the filaments grow slightly larger. More than 90% of purified platelet myosin molecules assemble into filaments in 0.1 M KC1 at pH 7. Purified preparations of the tail fragment of platelet myosin also form filaments. These filaments are slightly larger than myosin filaments formed under the same conditions, indicating that the size of the myosin filaments may be influenced by some interaction between the head and tail portions of myosin molecules. Calculations based on the size and shape of the myosin filaments, the dimensions of the myosin molecule and analysis of the bare zone reveal that the synthetic platelet myosin filaments consists of 28 myosin molecules arranged in a bipolar array with the heads of two myosin molecules projecting from the backbone of the filament at 14-15 nm intervals. The heads appear to be loosely attached to the backbone by a flexible portion of the myosin tail. Given the concentration of myosin in platelets and the number of myosin molecules per filament, very few of these thin myosin filaments should be present in a thin section of a platelet, even if all of the myosin molecules are aggregated into filaments.  相似文献   

16.
It has been reported that catch and striated muscle myosin heavy chains of scallop are generated through alternative splicing from a single gene [Nyitray et al. (1994) Proc. Natl. Acad. Sci. USA 91, 12686-12690]. They suggested that the catch muscle type myosin was expressed in various tissues of scallop, including the gonad, heart, foot, and mantle. However, there have been no reports of the primary structure of myosin from tissues other than the adductor muscles. In this study, we isolated a cDNA encoding the motor domain of myosin from the mantle tissue of scallop (Patinopecten yessoensis), and determined its nucleotide sequence. Sequence analysis revealed that mantle myosin exhibited 65% identity with Drosophila non muscle myosin, 60% with chicken gizzard smooth muscle myosin, and 44% with scallop striated muscle myosin. The mantle myosin has inserted sequences in the 27 kDa domain of the head region, and has a longer loop 1 structure than those of scallop striated and catch muscle myosins. Phylogenetic analysis suggested that the mantle myosin is classified as a smooth/nonmuscle type myosin. Western blot analysis with antibodies produced against the N-terminal region of the mantle myosin revealed that this myosin was specifically expressed in the mantle pallial cell layer consisting of nonmuscle cells. Our results show that mantle myosin is classified as a nonmuscle type myosin in scallop.  相似文献   

17.
P K Umeda  R Zak  M Rabinowitz 《Biochemistry》1980,19(9):1955-1965
Fast and slow myosin heavy chain mRNAs were isolated by indirect immunoprecipitation of polysomes from 14-day-old embryonic chick leg muscle. The antibodies were prepared against myosin heavy chains purified by NaDod-SO4-polyacrylamide gel electrophoresis and were shown to be specific for fast and slow myosin heavy chains. The RNA fractions directed the synthesis of myosin heavy chains in a cell-free translation system from wheat germ. Several smaller peptides were also synthesized in lower concentrations. These probably are partial products of myosin heavy chains, since they are immunoprecipitated with antibodies to myosin heavy chains. Immunoprecipitation of the translation products with the antibodies to fast and slow myosin heavy chains showed the RNA preparations to be approximately 94% enriched for fast myosin heavy chain mRNA and approximately 84% enriched for slow myosin heavy chain mRNA with respect to myosin HC type. Peptides having slightly different mobilities on NaDodSO4-polyacrylamide gels were immunoprecipitated by antibodies to fast and slow myosin heavy chains.  相似文献   

18.
Myosin rod was prepared from hen myosin by chymotryptic digestion. The indigested myosin was successfully removed by ultracentrifugation following alcohol treatment. No significant difference in UV absorption and CD spectra was observed between pH 7.0 and pH 10.5 for both myosin rod and myosin. When pH was raised to 11.7, the phenolic groups of the tyrosyl residues were ionized, and the helical configuration of the myosin rod and myosin could not withstand the electrostatic repulsion. When pH was further raised to 13.6, “abnormal” tyrosyl residues were ionized, resulting in decreased helix content. However, the myosin rod was stabler and less flexible against pH change than myosin, because of the lower content of tyrosyl residues in myosin rod.  相似文献   

19.
The myosin isozymes present in the developing rat soleus muscle from 1 week to 6 weeks after birth were investigated using biochemical and immunological methods. Electrophoresis of native myosin reveals that adult slow myosin is present in the soleus as early as 1 week after birth. At this time, embryonic and neonatal myosin can also be demonstrated. Using an immunotransfer technique, the presence of slow myosin heavy chain can be demonstrated at all time points examined whereas neonatal myosin heavy chain diminishes in quantity between 2 and 3 weeks, and is undetectable in the adult soleus. Specific polyclonal antibodies were prepared to embryonic, neonatal, and adult fast and slow myosins. Immunocytochemistry reveals a cellular heterogeneity at all stages examined. Different combinations of myosin isozymes can be found in the soleus fibers depending on the stage of development; these results suggest therefore that myosin isozyme transitions are occurring. Approximately half the fibers contain embryonic and slow myosin at 1 week after birth; these fibers subsequently contain only slow myosin. A second group of fibers contains embryonic and neonatal myosin at 1 week and most of them subsequently accumulate adult fast myosin. A portion of this latter group begins to acquire slow myosin from 4 weeks of age. These data are interpreted to suggest that a preprogrammed sequence of myosin isozymes is embryonic----neonatal----adult fast. At any time during development of an individual fiber, induction of slow myosin accumulation and repression of other types can occur.  相似文献   

20.
Embryonic chick heart ventricle myocytes retain the ability to alternate between proliferation and functional differentiation. A cytoplasmic isoform of myosin is present in cleavage furrows of various nonmuscle cells during cytokinesis, whereas one or more of the cardiac myosin isoforms are localized in sarcomeres of beating cardiomyocytes. Antibodies were employed to reveal the subcellular localizations of cytoplasmic and cardiac myosin isoforms in embryonic chick ventricle cardiomyocytes during cytokinesis. Monoclonal anticytoplasmic myosin antibodies were prepared against myosin purified from brains of 1-day-posthatched chickens and shown to react with chick brain myosin heavy chain by Western blots and/or ELISA tests. One monoclonal antibrain myosin antibody also cross-reacted with chick cardiac myosin but not with skeletal or smooth muscle myosins. Two antichick cardiac myosin monoclonal antibodies and one antichick skeletal myosin polyclonal antibody that cross-reacts with cardiac myosin were employed to identify cardiac sarcomeric myosin. Cells were isolated from day 8 embryonic chick heart ventricles, enriched for myocytes, grown in vitro for 3 days, and then examined by immunofluorescence techniques. Monoclonal antibodies against cytoplasmic myosin preferentially localized in the cleavage furrows of both cardiofibroblasts and cardiomyocytes in all stages of cytokinesis. In contrast, antibodies that recognize cardiac myosin were distributed throughout cardiomyocytes during early stages of cytokinesis, but became progressively excluded from the furrow area during middle and late stages of cytokinesis. These data suggest that in cells that contain both cytoplasmic and sarcomeric myosin isoforms, only cytoplasmic myosin isoforms are mobilized to from the contractile ring for cytokinesis.  相似文献   

设为首页 | 免责声明 | 关于勤云 | 加入收藏

Copyright©北京勤云科技发展有限公司  京ICP备09084417号