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C A Colaco

Publications and source records attributed to C A Colaco.

22 records · Page 2Linked to original sources
PubMed1983

Purification of the intermediate filament-associated protein, synemin, from chicken smooth muscle. Studies on its physicochemical properties, interaction with desmin, and phosphorylation.

Synemin, a 230,000-dalton protein associated with desmin- and vimentin-containing intermediate filaments (Granger, B. L., and Lazarides, E. (1980) Cell 22, 727-738), has been purified from gizzard smooth muscle and biochemically characterized. Purification was achieved by extracting the salt-insoluble pellet of muscle protein with 6 M urea and chromatography of the urea extract on columns of hydroxylapatite, DEAE-Sephacel, and phosphocellulose. The soluble form of synemin is a globular tetramer of 980,000 daltons with a S20,w of 22.4 +/- 3.2. Synemin has a pI of 5.34, in agreement with its high content in glutamic acid (20%), and is rich in serine (11%) and poor in cysteine (0.4%). Synemin is phosphorylated in smooth muscle and is one of the muscle proteins with the highest capacity to incorporate exogenously added [32P]phosphate. Of the [32P] phosphate incorporated into synemin, 95% is bound to serine and only 5% to threonine. The phosphorylation of synemin is enhanced by the cyclic AMP analog, 8-Br-cyclic AMP. Immunofluorescence studies using anti-synemin antibodies show that purified synemin binds to filaments of desmin assembled in vitro. Synemin specifically inhibits the immunoprecipitation of purified soluble desmin by anti-desmin antibodies, indicating that synemin interacts in vitro with soluble desmin.

Animals↗
PubMed1982

Partial purification of an intercalated disc-containing cardiac plasma membrane fraction.

The sarcolemma of cardiac muscle cells contains a specialised junctional region, the intercalated disc which includes three types of intercellular junction, the macula and fascia adherens and the nexus or gap junction. To facilitate the isolation of these junctions a procedure for the partial purification from mouse hearts of a subcellular fraction containing the intercalated disc region of the sarcolemma was developed. This involved investigating methods of tissue disruption that preserve the integrity of the intercalated disc and minimise myofibrillar entrapment of organelles. Examination of the distribution of marker enzymes showed that relative to the homogenate the intercalated disc fraction prepared by sucrose density centrifugation was only enriched 1.5- to 3-fold in 5'-nucleotidase and (Na+ + K+)-ATPase activities, whereas mitochondrial and sarcoplasmic reticulum marker enzymes were low. The properties of the intercalated disc-containing fraction were compared with the vesicular sarcolemmal fractions devoid of junctional complexes prepared by other methods.

5'-Nucleotidase↗
PubMed1981

A biochemical dissection of the cardiac intercalated disk: isolation of subcellular fractions containing fascia adherentes and gap junctions.

In view of our limited knowledge of the biochemical composition of intercellular junctions, a method was developed for the preparation from rats and mice of plasma membranes containing cardiac intercalated disks. When these membranes were extracted with detergents, e.g. N-lauryl sarcosinate or deoxycholate, the detergent-insoluble material contained structures derived mainly from fascia adherentes junctions, but a few gap junctions and maculae adherentes were also present. When the detergent extraction was carried out at an alkaline pH, the maculae adherentes junctions were dissolved. Fractionation of the detergent-insoluble extract on a sucrose gradient yielded a fraction containing fascia adherentes junction of density 1.20-1.26 g/cm3. Gap junctions banded at a lower density, 1.16-1.20 g/cm3. Polyacrylamide gel electrophoresis showed that the major polypeptide bands in the fascia adherentes-enriched fraction were of molecular weights 134000, 108000, 62-64000, 58000, 47000 and 43000. Although fractions with the gap junctions were contaminated by fascia adherentes junctions, the major polypeptides were calculated by subtraction to be of mol. wt 37000, 26000 and 19000. Two glycoproteins corresponding to minor polypeptides visualized by Coomassie Blue staining were present in the fascia adherentes fraction. Comparison of the fascia adherentes-enriched fraction with a Z-disc fraction prepared from rabbit hearts indicated a different morphology and polypeptide composition.

Animals↗