Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.11.18 (MAP)
 7,412 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Bovine myelin basic protein (MBP) was found to be an excellent in vitro substrate (apparent Km = 50 microM) for MAP (mitogen-activated protein) kinase and can be used in lieu of microtubule-associated protein 2 for purification and functional studies of the enzyme. MBP phosphotransferase activity co-purified with MAP kinase during sequential DE52, phenyl-Superose, and gel filtration chromatography, and kinase activities for the two substrates were co-regulated by mitogen stimulation. MAP kinase phosphorylated MBP exclusively on threonine, and only one major phosphopeptide was generated by digestion with trypsin or endoproteinase Lys-C. Using mass spectrometry, we determined that the phosphorylation site is threonine 97, present in the conserved triproline loop of MBP, with (partial) sequence -Thr-Pro-Arg-Thr97-Pro-Pro-Pro-. Thr97 is a known in vivo phosphorylation site in MBP although enzymes capable of phosphorylating this site have not been identified previously. MAP kinase phosphorylated peptide 88-109 from rabbit MBP and a synthetic peptide 91-109 from human MBP but did not phosphorylate either the histone H1 peptide, utilized by p34cdc2, or the peptide substrate for the recently described proline-directed kinase. Thus, the sequence surrounding threonine 97 in bovine MBP may contain essential features of a recognition sequence for MAP kinase.
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PMID:Identification by mass spectrometry of threonine 97 in bovine myelin basic protein as a specific phosphorylation site for mitogen-activated protein kinase. 170 Sep 79

The in vitro degradation of microtubule-associated protein 2 (MAP-2) and tubulin by the lysosomal aspartyl endopeptidase cathepsin D was studied. MAP-2 was very sensitive to cathepsin D-induced hydrolysis in a relatively broad, acidic pH range (3.0-5.0). However, at a pH value of 5.5, cathepsin D-mediated hydrolysis of MAP-2 was significantly reduced and at pH 6.0 only a small amount of MAP-2 was degraded at 60 min. Interestingly, the two electrophoretic forms of MAP-2 showed different sensitivities to cathepsin D-induced degradation, with MAP-2b being significantly more resistant to hydrolysis than MAP-2a. To our knowledge, this is the first clear demonstration that MAP-2 is a substrate in vitro for cathepsin D. In contrast to MAP-2, tubulin was relatively resistant to cathepsin D-induced hydrolysis. At pH 3.5 and an enzyme-to-substrate ratio of 1: 20, only 35% of the tubulin was degraded by cathepsin D at 60 min. The cathepsin D-mediated hydrolysis of tubulin was optimal only at pH 4.5. These results demonstrate that MAP-2 and tubulin are unequally susceptible to degradation by cathepsin D. These data also imply a potential for rapid degradation of MAP-2 in vivo by cathepsin D either in lysosomes or perhaps autophagic vacuoles of the neuron.
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PMID:Proteolysis of microtubule-associated protein 2 and tubulin by cathepsin D. 191 74

The in vitro degradation of microtubule-associated protein 2 (MAP-2) and spectrin by the calcium-dependent neutral protease calpain was studied. Five major results are reported. First, MAP-2 isolated from twice-cycled microtubules (2 X MT MAP-2) was extremely sensitive to calpain-induced hydrolysis. Even at an enzyme-to-substrate ratio (wt/wt) of 1:200, 2 X MT MAP-2 was significantly degraded by calpain. Second, MAP-2 purified from the total brain heat-stable fraction (total MAP-2) was significantly more resistant to calpain-induced hydrolysis compared with 2 X MT MAP-2. Third, MAP-2a and MAP-2b were proteolyzed similarly by calpain, although some relative resistance of MAP-2b was observed. Fourth, the presence of calmodulin significantly increased the extent of calpain-induced hydrolysis of the alpha-subunit of spectrin. Fifth, the two neuronal isoforms of brain spectrin (240/235 and 240/235E, referred to as alpha/beta N and alpha/beta E, respectively) showed different sensitivities to calpain. alpha N-spectrin was significantly more sensitive to calpain-induced degradation compared to alpha E-spectrin. Among other things, these results suggest a role for the calpain-induced degradation of MAP-2, as well as spectrin, in such physiological processes as alterations in synaptic efficacy, dendritic remodeling, and in pathological processes associated with neurodegeneration.
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PMID:Degradation of microtubule-associated protein 2 and brain spectrin by calpain: a comparative study. 201 58

An antibody directed against rat brain microtubule-associated protein 2 (MAP2) immunoprecipitated a protein of 240 kDa from a Xenopus oocyte extract. In contrast, in Xenopus brain extract, this antibody recognizes a protein of 280 kDa. The oocyte MAP2-related protein (called O-MAP) is present in both prophase I and metaphase II-blocked oocytes as demonstrated by immunoblotting experiments; it is in vivo phosphorylated. Immunocytochemical studies using the anti-rat brain MAP2 antibody demonstrated that the O-MAP colocalizes within the cortical microtubular array present in both prophase I and metaphase II oocytes. However, O-MAP is not associated with the microtubular structures which are organized during the oocyte prophase-metaphase transition, i.e., a giant cytoplasmic network and both the meiotic spindles. O-MAP therefore appears as a microtubule-associated protein oocyte specific and may play a role in the regulation of microtubule stability and the organization of the oocyte cytoskeleton.
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PMID:Intracellular localization of MAP2-related protein (O-map) in prophase I and metaphase II oocytes of Xenopus. 203 53

Several procedures were employed to examine the in vitro interaction between S-100 proteins and microtubule proteins. Binding of S-100 to tau factors was observed under all experimental conditions. S-100 binding to microtubule-associated protein 2 (MAP2) was best detected by exposing nitrocellulose-immobilized MAP2 or MAPs to either 125I-labeled S-100 or biotinylated S-100. S-100 binding to tubulin was detected when the two protein fractions were first incubated with each other followed by exposure to the bifunctional cross-linker disuccinimidylsuberate, and then separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and transfered onto nitrocellulose paper. By this procedure, complex formation between S-100 and tubulin, as well as between S-100 and a relatively low-molecular-weight MAP, was evidenced by immunoblotting using an anti-S-100 antiserum. Alternatively, complex formation between biotinylated S-100 and either tubulin or MAPs was visualized by means of avidin-peroxidase, after SDS-PAGE of the complex mixtures and transfer of the separated proteins onto nitrocellulose. The interaction between S-100 and tubulin was strictly Ca2+ dependent, and resistant to high concentrations of KCl, colchicine, or vinblastine.
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PMID:Molecular interaction of S-100 proteins with microtubule proteins in vitro. 266 85

Axoplasmic vesicles were purified and observed to translocate on isolated microtubules in an ATP-dependent, trypsin-sensitive manner, implying that ATP-binding polypeptides essential for force generation were present on the vesicle surface. To identify these proteins [alpha 32P]8-azidoadenosine 5'-triphosphate ([alpha 32P]8-N3ATP), a photoaffinity analogue of ATP, was used. The results presented here identify and characterize a vesicle-associated polypeptide having a relative molecular mass of 292 kD that bound [alpha 32P]8-N3ATP. The incorporation of label is ultraviolet light-dependent and ATP-sensitive. Moreover, the 292-kD polypeptide could be isolated in association with vesicles or microtubules, depending on the conditions used, and the data indicate that the 292-kD polypeptide is similar to mammalian brain microtubule-associated protein 2 (MAP 2) for the following reasons: The 292-kD polypeptide isolated from either squid axoplasm or optic lobe cross-reacts with antiserum to porcine brain MAP 2. Furthermore, it purifies with taxol-stabilized microtubules and is released with salt. Based on these characteristics, the 292-kD polypeptide is distinct from the known force-generating molecules myosin and flagellar dynein, as well as the 110-130-kD kinesin-like polypeptides that have recently been described (Brady, S. T., 1985, Nature (Lond.), 317:73-75; Vale, R. D., T. S. Reese, and M. P. Sheetz, 1985b, Cell, 42:39-50; Scholey, J. M., M. E. Porter, P. M. Grissom, and J. R. McIntosh, 1985, Nature (Lond.), 318:483-486). Because the 292-kD polypeptide binds ATP and is associated with vesicles that translocate on purified MAP-free microtubules in an ATP-dependent fashion, it is therefore believed to be involved in vesicle-microtubule interactions that promote organelle motility.
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PMID:Identification of a MAP 2-like ATP-binding protein associated with axoplasmic vesicles that translocate on isolated microtubules. 309 8

MAP2 (microtubule-associated protein 2) and tau factor are calmodulin-binding and actin filament-interacting proteins, respectively. We have examined the effect of Ca2+ and calmodulin on MAP-induced actin gelation by the low-shear falling-ball method, the high-speed centrifugation method, and electron microscopy using negative staining. Each MAP crosslinks actin filaments to increase the apparent viscosities and finally to form gels. Calmodulin inhibited MAP2- and tau factor-induced actin gelation (MAP2- and tau factor-actin interaction) only in the presence of Ca2+, but not in its absence. There were no differences in actin filament crosslinking activity of respective MAPs with or without Ca2+. MAP2 was not coprecipitated with F-actin only in the presence of Ca2+ and calmodulin determined by the high-speed centrifugation method. But MAP2 was found to bind to F-actin under any other conditions examined. In contrast, the tau factor-actin filament interaction could only be detected by the low-shear viscosity, but not by the high-speed centrifugation method. MAP2 and tau factor aggregated to form actin bundles as shown by electron microscopy. MAP2- or tau factor-induced bundle formation of actin filaments was inhibited only in the presence of Ca2+ and calmodulin, but not in the presence or absence of Ca2+. In conclusion, the interaction of MAP2- and tau factor-actin filaments is regulated by Ca2+ and calmodulin in a flip-flop switch.
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PMID:Ca2+ and calmodulin regulate microtubule-associated protein-actin filament interaction in a flip-flop switch. 400 97

Specific binding sites for the regulatory subunit of type II cAMP-dependent protein kinase (RII) were revealed in neurons by an immunohistochemical approach. Fixed frozen sections of several regions of the rat central nervous system were incubated in the presence of bovine RII. Bound bovine RII was subsequently detected by an immunofluorescence procedure using antibodies that recognize bovine but not rat RII. The results indicate that RII binds with high affinity to neurons. Binding is prominent in dendrites and almost undetectable in axons and axon terminals. The morphological distribution of the RII binding sites is almost identical to that of microtubule-associated protein 2 (MAP 2) immunoreactivity. Preadsorption of RII with a MAP preparation highly enriched in MAP 2 completely abolished binding of RII to tissue sections, suggesting that the binding is mediated by MAP 2. Our results indicate that frozen sections of fixed tissues are a suitable experimental system for study of specific interactions of cellular macromolecules at a morphological level.
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PMID:Frozen tissue sections as an experimental system to reveal specific binding sites for the regulatory subunit of type II cAMP-dependent protein kinase in neurons. 629 Oct 51

We used affinity-purified rabbit antibody to hog brain microtubule-associated protein 2 (MAP-2) to examine the pattern of attachment of MAPs to microtubules purified by cycles of in vitro assembly and disassembly. Microtubules were fixed, deposited on EM grids, and labeled with antibody and protein A-gold colloid followed by negative staining. We observed that: The sites of MAP attachment were greatly enhanced by antibody binding in negatively stained preparations. The axial repeat revealed by antibody (100 +/- 5 nm) was greater than the previously reported value of 32 nm based on thin sectioning and negative staining procedures. The antibody was arranged in a broad band and revealed a helical pattern of binding. Microtubules with and without treatment with alpha-chymotrypsin to remove the projection portion of MAP-2 looked similar, suggesting that the antibody-enhanced pattern may reflect the sites of MAP attachment on microtubules. Microtubules with an increased MAP:tubulin ratio exhibited the same 100-nm periodicity.
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PMID:The pattern of MAP-2 binding on microtubules: visual enhancement of MAP attachment sites by antibody labeling and electron microscopy. 667 2

The mechanism of stimulation of DNA synthesis by microtubule-associated protein 2 (MAP2) was examined in the nuclear matrix isolated from Physarum polycephalum. Porcine brain MAP2 stimulated DNA synthesis by the matrix with exogenous templates, but not with endogenous templates. Kinetic analyses showed that MAP2 decreases the Km of the matrix for deoxyribonucleoside triphosphates. Comparison of the Km values of active- and latent-type DNA replication machineries of Physarum suggested a possible role for MAPs or MAP-like proteins in DNA replication.
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PMID:Modulation of DNA synthesis by microtubule-associated protein 2 in the nuclear matrix isolated from Physarum polycephalum. 813 8


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