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Query: UMLS:C0272170 (SDS)
 50,377 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Two extremely high molecular weight proteins were found to be components of the intestinal epithelial cell brush border cytoskeleton. The largest brush border protein, designated T-protein, migrated on SDS gels as a doublet of polypeptides with molecular weights similar to muscle titin T I and T II. The other large brush border protein, designated N-protein, was found to have a polypeptide molecular weight similar to muscle nebulin. In Western analysis, a polyclonal antibody raised against brush border T-protein reacted specifically with T-protein in isolated brush borders and cross-reacted with titin in pectoralis and cardiac muscle samples. T-protein was distinguished from the muscle titins by an anti-cardiac titin mAb. A polyclonal antibody raised against N-protein was specific for N-protein in brush borders and cross-reacted with nothing in pectoralis muscle. Immunolocalization in cryosections of intestinal epithelia and SDS-PAGE analysis of fractionated brush borders revealed that both T-protein and N-protein are concentrated distinctly in the brush border terminal web region subjacent to the microvilli, but absent from the microvilli. EM of rotary-replicated T-protein samples revealed many of the molecules to be long (912 +/- 40 nm) and fibrous with a globular head on one end. In some of the molecules, the head domain appeared to be extended in a fibrous conformation yielding T-protein up to 1,700-nm long. The brush border N-protein was found as long polymers with a repeating structural unit of approximately 450 nm. Our findings indicate that brush border T-protein is a cellular isoform of titin and suggest that both T-protein and N-protein play structural roles in the brush border terminal web.
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PMID:Identification and characterization of two huge protein components of the brush border cytoskeleton: evidence for a cellular isoform of titin. 140 May 92

1. The post-mortem evolution of protein pattern in fish striated muscle was followed by SDS-PAGE, after different conditions of storage time and temperature. 2. Sarcoplasmic and sarcomeric fractions were analyzed respectively by low and high ionic strength extractions of fish muscle samples. 3. No evident modification of electrophoretic patterns was observed during the pre-rigor mortis period. 4. The high mol. wt proteins titin and nebulin were highly sensitive to proteolysis during the rigor mortis period. 5. Myosin extraction was predominantly influenced by the storage temperature. The myosin content of the extracts decreased during the rigor mortis period at storage temperatures greater than 8 degrees C. 6. alpha-Actinin was very resistant to proteolysis, but could be released from Z-disc structure during post-mortem aging.
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PMID:Sarcomeric disorganization in post-mortem fish muscles. 181 74

Ion-exchange column-purified I-protein was labeled by fluorescein isothiocyanate (FITC) at an equimolar ratio. When FITC-labeled I-protein was reacted with glycerinated myofibrils of chicken breast muscle in a phosphate-buffered saline, fluorescence was observed at the A-band and/or the Z-line of the sarcomere. However, FITC-labeled I-protein did not stain freshly prepared myofibrils. When FITC-I-protein was reacted with a nitrocellulose paper sheet on which muscle proteins were blotted after SDS-polyacrylamide gel electrophoresis, some peptide bands, including connectin and nebulin, were fluorescent. These facts can explain why anti-I-protein antibodies stain the A-I junctional region of fresh myofibrils and A-bands and/or Z-lines of glycerinated myofibrils. It is very likely that I-protein is transferred from the A-I junctions of myofibrils and translocates to A-bands and Z-lines, where some components that can bind to I-protein are localized, as myofibrils are degraded during the glycerination.
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PMID:FITC-labeled I-protein specifically binds to A-bands and/or Z-lines of glycerinated myofibrils of chicken breast muscle. 313 20

A wide range of phyla have been surveyed by SDS-PAGE for the new large proteins of the myofibril. Connectin (or titin) appears to be widely distributed. It is seen as a band of constant intensity and mobility in vertebrate striated muscle, but is absent from smooth muscle. It appears in more variable amounts, in a form of constant but greater mobility in many invertebrates: worms, molluscs (adductor but not gastropod feet), insects, a myriapod, and even in human blood platelets. Nebulin shares the same distribution in vertebrate muscles except for its notable absence in all heart muscle examined. It too is found in many invertebrates, not always with titin. It has been found in a worm, molluscs (adductor and gastropod feet), insects, crustaceans and an echinoderm. The mobility of nebulin varies within the vertebrates and more so between invertebrates (where, as with titin, it is greater). The isoforms of filamin in skeletal, cardiac, and smooth muscles of vertebrates are recorded. C-protein in rabbit muscles has four isoforms: white, alpha-red (X-protein), beta-red, and cardiac.
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PMID:A comparative study of high molecular weight proteins in various types of muscle across the animal kingdom. 375 64

An extensive network of transverse and longitudinal filamentous bridges was revealed when small myofibril bundles, prepared from Triton-EGTA-treated rabbit skeletal muscles, were extracted with Kl to remove the majority of thin and thick filaments. Transmission and scanning electron microscopic studies of these salt-resistant cytoskeletal residues indicated (a) small bundles of short transverse filaments connect adjacent myofibrils by forming Z to Z and M to M bridges; (b) parallel, continuous longitudinal filaments connect the peripheries of successive Z-disks and ensheath the sarcomere. These transverse and longitudinal filaments have the characteristic morphology of intermediate filaments; (c) two rings of tightly interwoven and tangled filaments, connected laterally by short filaments, encircle each Z disk. This double-ring also encircles a weblike meshwork which penetrates the sarcomeric space. From the peripheries of these rings, transverse and longitudinal intermediate filaments emerge; and (d) a massive amount of material translocated and accumulated near Z disks during Kl extraction. The residues were fairly resistant to solubilization by urea and SDS, and complete dissolution was achieved only with guanidinium chloride. SDS PAGE indicated that the residues consisted mainly of titin, nebulin, and variable amounts of residual myosin and actin. Desmin represented only a few percent of total residual proteins; however, it may be a major component of the intermediate filament network. We suggest that the intermediate filament should be considered an integral sarcomeric component that may play important cytoskeletal roles in muscle structure and mechanics.
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PMID:A network of transverse and longitudinal intermediate filaments is associated with sarcomeres of adult vertebrate skeletal muscle. 668 7

This study was conducted to determine degradation of the giant myofibrillar proteins titin and nebulin in postmortem aged beef, with known tenderness values, from animals differing in sex (steers vs bulls) and age (cows vs steers and bulls). Ten bulls and 10 steers (both groups were approximately 14 mo old) and 10 cows (44 to 108 mo old) were slaughtered. Longissimus muscle samples were removed for determination of Warner-Bratzler shear force, sensory panel tenderness evaluation, and SDS-PAGE analysis at 3, 7, 14, and 28 d postmortem. The SDS-PAGE analysis of titin and nebulin revealed that titin often migrated as three closely-spaced bands (T1, T1-2, T2, in increasing order of migration) in 3-d postmortem samples. With increasing time post-mortem, intact titin (T1) decreased and degraded titin (T2) increased in all samples. Within a class (i.e., steers, bulls, or cows) the rate of conversion of T1 to T2 was slower in the less-tender samples. The T1 to T2 conversion postmortem was slower in the intact males (bulls) than in the castrated males (steers). The T1 to T2 conversion postmortem also was slower in the older animals (cows) in comparison to the younger steers, but not in comparison to the younger bulls. Nebulin was degraded by 3 d postmortem in tender samples from steers, but some nebulin remained in the less-tender 3-d samples from steers and in all of the 3-d samples from bulls and older animals (cows). Intact nebulin was absent in all 7-d samples, regardless of the class of animal. Our results suggest that titin and nebulin are degraded at faster rates in more tender beef samples within each of the three classes of animals examined. The rate of degradation seems to differ when sex and age classifications are compared.
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PMID:Effects of postmortem aging time, animal age, and sex on degradation of titin and nebulin in bovine longissimus muscle. 762 49

Changes in the molecular types of connectin and nebulin during development of chicken breast and leg muscles were determined by an improved SDS-polyacrylamide gel electrophoresis (PAGE) using 2% polyacrylamide slab gel. The adult leg-type alpha-connectin (alpha L-connectin) and nebulin (L-nebulin) appeared in embryonic breast muscle, and changed into the adult breast-type ones (alpha B-connectin, B-nebulin) specific for adult breast muscle after hatching. In leg muscle, alpha L-connectin and L-nebulin appeared in an embryonic stage, and remained unchanged in molecular types throughout the entire process of development. alpha-Connectin and nebulin seemed to be regulated by a similar mechanism during development. On the other hand, beta-connectin appeared in an earlier stage of development of the embryonic breast muscle, independently of alpha-connectin.
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PMID:Changes in the molecular types of connectin and nebulin during development of chicken skeletal muscle. 776 56

Nebulin, a giant protein (molecular mass 800 kDa) specific for the skeletal muscle of vertebrates, has been suggested to be involved in the length regulation of the thin filament as a 'molecular ruler'. Despite its size, nebulin appears to be composed mainly of small repeats of approximately 35 amino acids. We have characterized in this study the conformational and functional properties of single repeats. Complete repeats were found to bind to F-actin while a truncated one did not. One repeat is therefore the smallest unit for nebulin--actin interaction. Circular dichroism and nuclear magnetic resonance spectra measured for the peptides in water indicated a transient helical conformation. The folded region is located for them all around the conserved sequence SDxxYK. The helical conformation is strongly stabilized by anionic detergents and trifluoroethanol while uncharged or positively charged detergents have no effect. Since the surface of the actin filament is known to contain clusters of negative charges, anionic detergents may mimic the effect of an actin environment. 3D structures were calculated for three representative peptides in SDS. In vivo, the nebulin helices should form a complex with the actin filament. Based on the assumed importance of charge interactions between nebulin and actin, we propose a model for the structure of the F-actin-nebulin complex in vivo. According to that, two nebulin molecules occupy symmetrical positions along the central cleft of the actin filament bridging the two strands of the actin two-start helix. The consistency of this model with experimental data is discussed.
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PMID:Nebulin, a helical actin binding protein. 816 78

Postmortem (PM) and mu-calpain-induced degradation of specific skeletal muscle proteins was monitored by SDS-PAGE and Western blotting. Samples were removed from bovine longissimus thoracis (LT) at approximately 45 min PM for the preparation of at-death (0-d) myofibrils (MF). The LT was excised at 1 d PM, vacuum-packaged, and stored at 2 degrees C. Samples were removed for Warner-Bratzler shear force analysis and biochemical analysis at 1, 3, 7, 14, 28, and 56 d PM. The protease mu-calpain was purified from bovine skeletal muscle and used to digest at-death MF at pH 5.6, 4 degrees C, 100 microM CaCl2. Degradation of the proteins titin, nebulin, filamin, desmin, and troponin-T was monitored in the PM and mu-calpain-digested samples by using SDS-PAGE and Western blotting. The PM samples that had significantly lower shear force (LSF) values (P < .05) at 1 d PM exhibited faster degradation of these five proteins than the higher shear force (HSF) samples. In LSF samples, the intact titin band (T1) was absent by 7 d PM and nebulin was absent by 3 d PM. In LSF samples, some filamin was degraded by 3 d PM, but in HSF samples degradation was not apparent until 14 d PM. In LSF samples, desmin was degraded more rapidly PM than in HSF samples. Troponin-T was broken down PM to yield two major polypeptides of approximately 28 and 30 kDa; these polypeptides appeared earlier PM in LSF samples. Degradation products, similar to those observed PM, for all five proteins also were detected in Western blots of mu-calpain-digested MF, suggesting the calpain system plays a key role in PM protein degradation.
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PMID:Proteolysis of specific muscle structural proteins by mu-calpain at low pH and temperature is similar to degradation in postmortem bovine muscle. 872 31

Purified myofibril (MF) and homogenized whole muscle (WM) samples were prepared from A maturity market steers. Samples were removed at 0, 1, 3, 7, 14, and 28 d postmortem. The MF and WM samples from all steers were analyzed by SDS-PAGE (5% gels) and by Western blot analysis using monoclonal antibodies to titin and nebulin. The rates of degradation of the intact forms of titin and nebulin, with regard to differences dependent on sample type (MF vs WM), were examined. The results showed that there was very little difference in the rate of postmortem degradation of the intact form of titin or of intact nebulin with respect to the two types of samples examined. Analysis of MF and WM preparations revealed that titin and nebulin were progressively degraded, each at its own rate, with nebulin degrading faster, as postmortem storage time increased. Examination of MF and WM samples showed that the intact form of titin (T1) was absent at the same time postmortem in both sample types. Intact nebulin was not detected in MF and WM preparations at the same time postmortem with respect to sample type examined. Our results indicate that either purified MF or WM samples can be used satisfactorily to analyze the rate of degradation of the intact forms of both titin and nebulin.
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PMID:Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and western blotting comparisons of purified myofibrils and whole muscle preparations for evaluating titin and nebulin in postmortem bovine muscle. 872 98


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