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11.
Hans Georg B?umert Akitsugu Kenmoku Gert Middelhoff Franz Ortanderl Alexander Thrun Heinz Faulstich Wolfgang Schiebler Hugo Fasold 《Journal of Protein Chemistry》1988,7(5):571-580
With the aid of tartryl-bis--aminocaprylazide artificial dimers were produced from F actin from rabbit striated muscle. These derivatives will not polymerize by themselves but are able to copolymerize fully with native G actin. By modification of a single side chain per dimer, this copolymerization was completely inhibited. The dimers are able to activate subfragment I ATPase of myosin and bind to DNase I with inactivation of the enzyme in the same manner as native G actin. Within the dimer, one ADP is immobilized and will exchange against ATP extremely slowly. The dimers do not bind to the mushroom toxin phalloidin. 相似文献
12.
The recent discovery of DNA sequences responsible for the specific attachment of chromosomal DNA to the nuclear skeleton (MARs/SARs) was an important step towards our understanding of the functional and structural organization of eukaryotic chromatin [Mirkovitch et al.: Cell 44:273-282, 1984; Cockerill and Garrard: Cell 44:273-282, 1986]. A most important question, however, remains the nature of the matrix proteins involved in the specific binding of the MARs. It has been shown that topoisomerase II and histone H1 were capable of a specific interaction with SARs by the formation of precipitable complexes [Adachi et al.: EMBO J8:3997-4006, 1989; Izaurralde et al.: J Mol Biol 210:573-585, 1989]. Here, applying a different approach, we were able to "visualize" some of the skeletal proteins recognizing and specifically binding MAR-sequences. It is shown that the major matrix proteins are practically the same in both salt- and LIS-extracted matrices. However, the relative MAR-binding activity of the individual protein components may be different, depending on the method of matrix preparation. The immunological approach applied here allowed us to identify some of the individual MAR-binding matrix proteins. Histone H1 and nuclear actin are shown to be not only important components of the matrix, but to be involved in a highly efficient interaction with MAR-sequences as well. Evidence is presented that proteins recognized by the anti-HMG antibodies also participate in MAR-interactions. 相似文献
13.
The accessibility of etheno-nucleotides to collisional quenchers and the nucleotide cleft in G- and F-actin.
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Recent publication of the atomic structure of G-actin (Kabsch, W., Mannherz, H. G., Suck, D., Pai, E. F., & Holmes, K. C., 1990, Nature 347, 37-44) raises questions about how the conformation of actin changes upon its polymerization. In this work, the effects of various quenchers of etheno-nucleotides bound to G- and F-actin were examined in order to assess polymerization-related changes in the nucleotide phosphate site. The Mg(2+)-induced polymerization of actin quenched the fluorescence of the etheno-nucleotides by approximately 20% simultaneously with the increase in light scattering by actin. A conformational change at the nucleotide binding site was also indicated by greater accessibility of F-actin than G-actin to positively, negatively, and neutrally charged collisional quenchers. The difference in accessibility between G- and F-actin was greatest for I-, indicating that the environment of the etheno group is more positively charged in the polymerized form of actin. Based on calculations of the change in electric potential of the environment of the etheno group, specific polymerization-related movements of charged residues in the atomic structure of G-actin are suggested. The binding of S-1 to epsilon-ATP-G-actin increased the accessibility of the etheno group to I- even over that in Mg(2+)-polymerized actin. The quenching of the etheno group by nitromethane was, however, unaffected by the binding of S-1 to actin. Thus, the binding of S-1 induces conformational changes in the cleft region of actin that are different from those caused by Mg2+ polymerization of actin.(ABSTRACT TRUNCATED AT 250 WORDS) 相似文献
14.
Q Q Zhuang S Rosenberg J Lawrence A Stracher 《Biochemical and biophysical research communications》1984,118(2):508-513
Actin binding protein from human blood platelets is shown to exist in the resting platelet as a phosphorylated protein and contains two residues of phosphate per 260,000 kd. Removal of one-half of these residues with E. coli alkaline phosphatase results in the loss of its ability to crosslink F-actin into a low speed sedimentable complex (its cytoskeleton) and to bind to an F-actin affinity column. Thus, phosphorylation-dephosphorylation of ABP may be an important regulatory mechanism by which the platelet regulates its shape via its cytoskeletal structure. 相似文献
15.
Y Gache F Landon H Touitou A Olomucki 《Biochemical and biophysical research communications》1984,124(3):877-881
Purified alpha-actinin from human platelets was digested with Ca2+-activated protease from muscle. The alpha subunit (Mr = 100 kDa) was degraded into a unique polypeptide b of slightly lower molecular mass. In fresh platelets, only the a subunit was detected by immunoblotting techniques, while in out-dated platelets, both a and b polypeptides were present. Since a similar conversion of a to b occurs in vitro as in whole platelets, it can be assumed that, in platelets, alpha-actinin is cleaved by the endogenous Ca2+-activated protease. 相似文献
16.
The sperm head of the plains mouse, Pseudomys australis, has three curved hooks projecting from its anterior margin. The two ventral hooks have previously been shown to consist largely of an extension of the subacrosomal material. To characterize further the structure and composition of the ventral hooks, we have examined their formation during spermiogenesis using transmission electron microscopy, silver staining, and actin localization with NBD-phallacidin. The ventral hooks develop as an extension of the perinuclear space and postacrosomal dense lamina on the anteroventral margin of the sperm head. Bundles of 6-nm-thick filaments appear in the core of each hook; these are probably actin filaments based on staining of the hooks with NBD-phallacidin. Just prior to spermiation, electron-dense material condenses in the core of the ventral hooks and concurrently in the perinuclear space in the remainder of the sperm head. The two ventral hooks thus appear to consist of a core of perinuclear material and actin filaments, which is enclosed by a continuation of the postacrosomal dense lamina. 相似文献
17.
C. Méjean M. Boyer J. P. Labbé J. Derancourt Y. Benyamin C. Roustan 《Bioscience reports》1986,6(5):493-499
The interaction of two different anti-actin antibody populations with the myosin subfragment 1-F-actin rigor complex has been studied. In contrast with the 1–7 sequence, the 18–28 sequence appears to be strongly implicated in the contact area of the myosin head on the actin polypeptide chain. 相似文献
18.
T D Pollard 《Journal of cellular biochemistry》1986,31(2):87-95
19.
Human arterial smooth muscle cells in culture: Inverse relationship between proliferation and expression of contractile proteins 总被引:5,自引:0,他引:5
Gunnar Fager Göran K. Hansson Allen M. Gown David M. Larson Omar Skalli Göran Bondjers 《In vitro cellular & developmental biology. Plant》1989,25(6):511-520
Summary Human arterial smooth muscle cells (hASMC) from explants of the inner media of uterine arteries were studied in secondary
culture. We had previously found that these cells depend on exogenous platelet-derived growth factor (PDGF) for proliferation
in vitro. Deprivation of the serum mitogen(s) by culture in plasma-derived serum or bovine serum albumin (BSA) caused a true
growth arrest that was reversible upon reexposure to the mitogen(s). When added to serum-containing medium, heparin caused
a reversible growth arrest which could be competed for by increasing concentrations of serum. In the current study we used
a set of smooth muscle-specific actin and myosin, antibodies to study the expression of contractile proteins in stress fibers
under indirect immunofluorescence on hASMC in culture. Even in sparse culture, grwoth-arrested hASMC expressed stress fibers
containing these actin and myosin epitopes. This was true irrespective of whether growth arrest was achieved by culture in
media containing only BSA or a combination of heparin and whole blood serum. hASMC proliferating in whole blood serum in sparse
culture did not express such strees fibers, as judged by immunofluorescent staining. This was true also for cells that were
restimulated to proliferate in serum after a growth arrest. Utilizing a monoclonal antibody against a nuclear antigen expressed
in proliferating human cells, we were able to demonstrate an inverse relationship between the expression of this antigen and
the SMC-specific contractile proteins, respectively. Under these culture conditions, the reversible transition between defifferentiated
and differentiated hASMC was almost complete and terminated about 1 wk after the change in culture condition. We conclude
that hASMC in vitro respond, to exogenous PDGF by proliferation and dedifferetiation as a single population of cells. We also
conclude that this modulation is reversible, because the cells become uniformly quiescent and differentiated when the mitogenic
stimulus is blocked or removed.
This study was supported by grants from the Swedish Medical Research Council (Project no. 4531 and 6816), the Swedish Association
against Heart and Chest Diseases, the King Gustaf V and Queen Victoria Foundation, the National Institutes of Health, Bethesda,
MD (grant HL 29873) and the Swedish National Board for Laboratory Animals. 相似文献
20.
Summary Alpha-smooth muscle actin is currently considered a marker of smooth muscle cell differentiation. However, during various
physiologic and pathologic conditions, it can be expressed, sometimes only transiently, in a variety of other cell types,
such as cardiac and skeletal muscle cells, as well as in nonmuscle cells. In this report, the expression of actin mRNAs in
cultured rat capillary endothelial cells (RFCs) and aortic smooth muscle cells (SMCs) has been studied by Northern hybridization
in two-dimensional cultures seeded on individual extracellular matrix proteins and in three-dimensional type I collagen gels.
In two-dimensional cultures, in addition to cytoplasmic actin mRNAs which are normally found in endothelial cell populations,
RFCs expressed α-smooth muscle (SM) actin mRNA at low levels. α-SM actin mRNA expression is dramatically enhanced by TGF-β1. In addition, double immunofluorescence staining with anti-vWF and anti-α-SM-1 (a monoclonal antibody to α-SM actin) shows
that RFCs co-express the two proteins. In three dimensional cultures, RFCs still expressed vWF, but lost staining for α-SM
actin, whereas α-SM actin mRNA became barely detectable. In contrast to two-dimensional cultures, the addition of TGF-β1 to the culture media did not enhance α-SM actin mRNA in three-dimensional cultures, whereas it induced rapid capillary tube
formation. Actin mRNA expression was modulated in SMCs by extracellular matrix components and TGF-β1 with a pattern very different from that of RFCs. Namely, the comparison of RFCs with other cell types such as bovine aortic
endothelial cells shows that co-expression of endothelial and smooth muscle cell markers is very unique to RFCs and occurs
only in particular culture conditions. This could be related to the capacity of these microvascular endothelial cells to modulate
their phenotype in physiologic and pathologic conditions, particularly during angiogenesis, and could reflect different embryologic
origins for endothelial cell populations.
Supported by a Post-Doctoral Fellowship from the Swiss National Science Foundation (OK) and grant HL-RO1-28373 (JAM) from
the Department of Human Services, Public Health Service, Washington, D.C. 相似文献