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Query: UNIPROT:Q07644 (polypeptide)
 72,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The membrane polypeptides of growth cone fragments ("growth cone particles," GCPs) isolated from fetal rat brain by subcellular fractionation have been analyzed in further detail. The major polypeptides of salt-washed GCP membranes detected by 1-dimensional gel electrophoresis (Ellis et al., 1985b) resolve in 2-dimensional gels as a spot of 52 kDa that comigrates with beta-tubulin and reacts with anti-beta-tubulin; a 46 kDa, pl 4.3, polypeptide (pp46) that has no equivalent in the soluble fraction and is identical to one of the GCP's major phosphoproteins (Katz et al., 1985) and to GAP43 (Willard et al., 1985); a spot of 42 kDa that comigrates with actin; and a species of 34 kDa (p34) without soluble equivalent. The prominent 38 kDa doublet identified in 1-dimensional gels is difficult to resolve in 2-dimensional gels. The major phosphoproteins pp80ac, pp46, and pp40 (Katz et al., 1985), as well as p34 partition into the oil phase of Triton X-114 extracts, suggesting that they are integral membrane proteins, at least in our experimental conditions. The properties of pp46 reported here are in conflict with the highly hydrophilic amino acid sequence predicted for GAP43/B50/F1 (Basi et al., 1987; Karns et al., 1987). Growth-cone and presynaptic membrane proteins are compared as follows. After eye injection of 35S-methionine, GCPs and synaptosomes are isolated from the target areas of optic nerve of fetal and adult rats, respectively. Polypeptides are separated by 1- and 2-dimensional gel electrophoresis and the radiolabeled species identified fluorographically. The comparison of labeled GCP and synaptosome polypeptides shows that all 5 major Coomassie blue-stained polypeptides of GCP membranes (52, 46, 42, 38, 34 kDa) are intensely labeled after eye injection. However, in synaptosomes, these polypeptides are weakly labeled if at all; instead, an intensely labeled polypeptide of 28 kDa, and several additional species not seen in GCPs, have appeared. Therefore, the major growth cone membrane proteins are developmentally regulated, and the rates of synthesis and transport into the axonal ending of neuronal polypeptides change dramatically at the time of synaptogenesis.
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PMID:Membrane proteins of the nerve growth cone and their developmental regulation. 292 76

Aspergillus nidulans has two beta-tubulin genes: benA, which is involved in both vegetative growth and asexual sporulation, and tubC, which is involved mainly in asexual sporulation. Both genes have now been cloned and sequenced. benA encodes a polypeptide of 447 amino acids (aa) and tubC encodes one of 449 aa. The two polypeptides differ by 78 aa residues but the net charge for the two proteins remains the same. The divergence between the amino acid sequences of the Aspergillus beta-tubulins is greater than that for any other two beta-tubulins yet described in the same organism. The benA gene has eight introns and the tubC gene has five, all of which correspond in amino acid position to introns in benA. The positions of some of these introns are conserved in other beta-tubulin genes. The 5'-splice site, internal, and 3'-splice site consensus sequences are similar to those found in other fungal introns. The transcriptional start points for each gene have been determined using primer extension and/or S1 nuclease mapping. Neither the benA gene nor the tubC gene contains a TATA sequence in its 5'-flanking region. The tubC gene has two repeated upstream sequences which are not found in benA. The sites of polyadenylation have been determined for each gene using S1 nuclease mapping. Neither gene contains a polyadenylation signal, AATAAA, typical of other eukaryotic genes.
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PMID:Aspergillus nidulans beta-tubulin genes are unusually divergent. 295 91

We have determined the nucleotide sequence of the chicken beta 5 (c beta 5)-tubulin gene. The gene displayed the coding structure common to all previously studied vertebrate beta-tubulin genes and was divided into four exon sequences interrupted by three intervening sequences (located between codons 19 and 20, within codon 56, and within codon 93). Comparison of the predicted polypeptide sequence encoded by c beta 5 with those of four other available chicken beta-tubulin sequences revealed that c beta 5 encoded a highly divergent beta-tubulin polypeptide isotype which was distinguished from previously known sequences primarily by two discrete variable sequence domains. However, c beta 5 uniquely shared identity in 16 residue positions with another divergent chicken beta-tubulin gene, c beta 4. These common sequences distinguished c beta 4 and c beta 5 from the remaining three chicken beta-tubulin genes. Analysis of the expression of c beta 5 and c beta 4 revealed a strikingly complementary pattern of gene expression: c beta 5 was expressed in a wide variety of cell and tissue types but not in neurons, whereas c beta 4 expression was detected uniquely in neuronal cells. Overall, these findings suggest the existence of two divergent families of beta-tubulin sequences in the chicken and further raise the possibility that the complementary expression of the c beta 4 and c beta 5 genes may fulfill a requirement for the presence of a divergent beta-tubulin polypeptide isotype in all cell types.
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PMID:Sequence and expression of the chicken beta 5- and beta 4-tubulin genes define a pair of divergent beta-tubulins with complementary patterns of expression. 302 56

beta-tubulin of budding yeast Saccharomyces cerevisiae is a polypeptide of 457 amino acids encoded by the unique gene TUB2. We investigated the function of the carboxy-terminal part of yeast beta-tubulin corresponding to the carboxy-terminal variable domain of mammalian and avian beta-tubulins. The GAA codon for Glu-431 of TUB2 was altered to TAA termination codon by using in vitro site-directed mutagenesis so that the 27-amino acid residues of the carboxyl terminus was truncated when expressed. The mutagenized TUB2 gene (tub2(T430)) was introduced into a haploid strain in which the original TUB2 gene had been disrupted. The tub2(T430) haploid strain grows normally less than 30 but not at 37 degrees C. The truncation of the carboxyl terminus caused hypersensitivity to antimitotic drugs and low spore viability at the permissive temperature for vegetative growth. Immunofluorescence labeling with antitubulin antibody and DNA staining with 4',6'-diamidino-2-phenylindole showed that in these cells at 37 degrees C, formation of spindle microtubules and nuclear division was inhibited and cytoplasmic microtubule distribution was aberrant. These results suggest that functions of the carboxy-terminal domain of yeast beta-tubulin are necessary for cells growing under suboptimal growth conditions although it is not essential for growth under the optimal growth conditions. Cells bearing tub2(411), a tub2 gene in which the GAA codon for Glu-412 was altered to TAA were no more viable at any temperature. In addition, a haploid strain carrying two functional beta-tubulin genes is not viable.
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PMID:Truncation of the carboxy-terminal domain of yeast beta-tubulin causes temperature-sensitive growth and hypersensitivity to antimitotic drugs. 304 21

The transition from the promastigote stage to the amastigote stage in Leishmania appears to involve a sequence of steps which enable the parasite to adapt to its new environment. In this study, transformation from the promastigote to an amastigote-like stage was induced by temperature elevation and the effects on protein synthesis and the mRNA population were analyzed. Whereas significant changes in the polypeptide complement of the cell were observed, few, if any, changes were seen at the level of the mRNAs as determined by translation in a cell-free system. Increasing the growth temperature caused a rapid cessation of beta-tubulin synthesis but the abundance and sizes of mRNAs specific for beta-tubulin were unaltered. These data suggest that the regulation of beta-tubulin under these conditions is occurring at the level of translation.
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PMID:Effects of temperature elevation on mRNA and protein synthesis in Leishmania mexicana amazonensis. 317 31

beta-Tubulin is encoded in the genomes of higher animals by a small multigene family comprising approximately seven functional genes. These genes produce a family of closely related, but distinct polypeptide isotypes that are distinguished principally by sequences within the approximately 15 carboxy-terminal amino acid residues. By immunizing rabbits with chemically synthesized peptides corresponding to these variable domain sequences, we have now prepared polyclonal antibodies specific for each of six distinct isotypes. Specificity of each antiserum has been demonstrated unambiguously by antibody binding to bacterially produced, cloned proteins representing each isotype and by the inhibition of such binding by preincubation of each antiserum only with the immunizing peptide and not with heterologous peptides. Protein blotting of known amounts of cloned, isotypically pure polypeptides has permitted accurate quantitative measurement of the amount of each beta-tubulin isotype present in the soluble and polymer forms in various cells, but has not revealed a bias for preferential assembly of any isotype. Localization of each isotype in three different cell types using indirect immunofluorescence has demonstrated that in vivo each class of microtubules distinguishable by light microscopy is assembled as copolymers of all isotypes expressed in a single cell.
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PMID:In vivo microtubules are copolymers of available beta-tubulin isotypes: localization of each of six vertebrate beta-tubulin isotypes using polyclonal antibodies elicited by synthetic peptide antigens. 331 37

alpha- and beta-Tubulin are encoded in vertebrate genomes by a family of approximately 6-7 functional genes whose polypeptide products differ in amino acid sequence. In the chicken, one beta-tubulin isotype (c beta 6) has previously been found to be expressed only in thrombocytes and erythroid cells, where it is assembled into a circumferential ring of marginal band microtubules. In light of its unique in vivo utilization and its divergent assembly properties in vitro, we used DNA transfection to test whether this isotype could be assembled in vivo into microtubules of divergent functions. Using an antibody specific to c beta 6, we have found that upon transfection this polypeptide is freely coassembled into an extensive array of interphase cytoplasmic microtubules and into astral and pole-to-chromosome or pole-to-pole microtubules during mitosis. Further, examination of developing chicken erythrocytes reveals that both beta-tubulins that are expressed in these cells (c beta 6 and c beta 3) are found as co-polymers of the two isoforms. These results, in conjunction with efforts that have localized various other beta-tubulin isotypes, demonstrate that to the resolution limit afforded by light microscopy in vivo microtubules in vertebrates are random copolymers of available isotypes. Although these findings are consistent with functional interchangeability of beta-tubulin isotypes, we have also found that in vivo microtubules enriched in c beta 3 polypeptides are more sensitive to cold depolymerization than those enriched in c beta 6. This differential quantitative utilization of the two endogenous isotypes documents that some in vivo functional differences between isotypes do exist.
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PMID:In vivo coassembly of a divergent beta-tubulin subunit (c beta 6) into microtubules of different function. 331 49

Membrane elements in brain tissue contain relatively large amounts of alpha- and beta-tubulin (FIGURES 2 and 3). We have investigated the subcellular sites of tubulin biosynthesis in order to determine the origin of this membrane-associated tubulin. Free and membrane-bound polysomes from rat forebrain were separated by differential centrifugation, and the products of translation from these polysome populations were analyzed by 2DGE (FIGURES 4 and 6). Alpha- and beta-tubulin subunits were synthesized by the free polysome population (FIGURES 4 and 5A and B). The membrane-bound polysome fraction synthesized a protein with similar (but not identical) characteristics to alpha-tubulin (denoted as "MB" in FIGURE 6), including isoelectric point, molecular weight, peptide map, and copurification with microtubules after aggregation-disaggregation. Tubulin subunits synthesized in vitro by free polysomes could associate posttranslationally with a microsome fraction (FIGURE 7A). The association of the tubulin translation products with membranes was not disrupted by high salt; the associated tubulin, however, was susceptible to proteolytic digestion, with the exception of one of the beta-tubulin subunits (FIGURE 7B). There was an identical protease-resistant beta-tubulin subunit among the native proteins of the smooth microsome fractions. Our data is consistent with the conclusion that at least one beta subunit of membrane-associated tubulin is synthesized by free polysomes and becomes posttranslationally added to membrane structures. It is unlikely that a cotranslational mechanism is responsible, in which there is a signal-mediated insertion of a growing polypeptide chain to membrane. Our results, however, are consistent with a "membrane trigger" mechanism proposed by Wickner in which the membrane lipid bilayer triggers the folding of a polypeptide into a configuration that allows integral membrane insertion. The association of tubulin with membranes may also be secondary to the interaction of hydrophobic elements. The amino acid sequence of beta tubulin is known to contain several hydrophobic domains. Tubulin can be incorporated into phospholipid vesicles and various subcellular membrane elements. In our studies, in vitro synthesized tubulin from free polysome was found to be purified by hydrophobic affinity chromatography with ethane-sepharose (FIGURE 8). Thus, the hydrophobic characteristics of newly synthesized tubulin could be partially responsible for the posttranslational association of tubulin subunit with membranes. Native tubulin in a soluble fraction of CNS tissue was not purified by hydrophobic affinity chromatography.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:In vitro studies of the biosynthesis of brain tubulin on membranes. 346 Apr 61

A radioactive, photoactive Vinca alkaloid, N-(p-azido-[3,5-3H]-benzoyl)-N'-beta-aminoethylvindesine [( 3H]NABV) with pharmacological and biological activities similar to vinblastine was synthesized and used to identify specific Vinca alkaloid macromolecular interactions in calf brain homogenate by photoaffinity labeling. The most prominent photolabeled species were 54.3- and 21.5-kDa polypeptides. The Vinca alkaloid-binding specificity of these polypeptides was confirmed by competitive blocking of specific photolabeling by vinblastine but not by colchicine or daunorubicin. The 54.3- and 21.5-kDa polypeptides exhibited specific half-maximum saturable photolabeling at 2.1 and 0.95 X 10(-7) M [3H]NABV, respectively. Relative vinblastine and NABV association constants (Ka vinblastine/Ka NABV) for the 54.3- and 21.5-kDa polypeptides were estimated to be 0.86 and 1.4, respectively. The 54.3-kDa component was found in both high speed (100,000 X g; 1 h) pellet and supernatant fractions, whereas the 21.5-kDa component was located primarily in the high speed pellet. Photolabeling of both components was maximal after 12-min UV light exposure, linear up to 120 micrograms of homogenate protein and only slightly affected by the nitrene scavenger p-aminobenzoic acid. The 54.3-kDa polypeptides of [3H]NABV-photolabeled calf brain high speed supernatant and detergent-solubilized high speed pellet fractions were identified as tubulin subunits by immunoprecipitation with monoclonal antibodies to alpha- or beta-tubulin subunits. Although the identity and function of the 21.5-kDa polypeptide is not known, this polypeptide may have a role in membrane-related effects of the Vinca alkaloids. These results demonstrate that [3H]NABV is an attractive tool for identifying and characterizing specific high affinity vinblastine cellular polypeptide acceptors which may initiate or mediate known and unknown mechanisms of Vinca alkaloid action.
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PMID:Specific Vinca alkaloid-binding polypeptides identified in calf brain by photoaffinity labeling. 354 1

We report the determination of the complete DNA sequence for c beta 3, a chicken beta-tubulin gene which we show to be the dominant beta-tubulin expressed in testis. Like all previously studied vertebrate beta-tubulin genes, the gene is divided into four exon sequences interrupted by three intervening sequences (located between amino acids 19 and 20, within codon 56, and within codon 93). Analysis of the program of expression of this gene indicates that it encodes the dominant chicken testis beta-tubulin, although it is also expressed at lower levels in a wide variety of cell and tissue types. Comparison of the predicted polypeptide sequence for c beta 3 with four other available chicken beta-tubulin genes confirms our earlier suggestion that within an otherwise conserved framework, sequences within two variable region domains serve to define specific beta-tubulin polypeptide isotypes. The data indicate that the c beta 3 gene encodes a unique beta-tubulin isotype which diverges from the dominant neuronal beta-tubulin isotype in 18 of 445 residues (4%). Although the protein coding regions of the c beta 3 gene are highly homologous to the chicken c beta 1, c beta 2, c beta 4, and c beta 5 genes previously reported by us, no significant sequence homology with these previously analyzed genes is discernible in the 5'- or 3'-untranslated region sequences, in the intervening sequences, or in the presumptive transcriptional promoter sequences.
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PMID:Sequence and expression of the chicken beta 3 tubulin gene. A vertebrate testis beta-tubulin isotype. 375 66


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