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Biomedical subjects

S R Karr

Publications and source records attributed to S R Karr.

5 recordsLinked to original sources
PubMed1988

The synthesis of murine ferrochelatase in vitro and in vivo.

Ferrochelatase (protohaem ferro-lyase, EC 4.99.1.1), the terminal enzyme of the haem-biosynthetic pathway, is an integral membrane protein of the mitochondrial inner membrane. When murine erythroleukaemia cells are labelled in vivo with [35S]methionine, lysed, and the extract is immunoprecipitated with rabbit anti-(mouse ferrochelatase) antibody, a protein of Mr 40,000 is isolated. However, when isolated mouse RNA is translated in a cell-free reticulocyte extract, a protein of Mr 43,000 is isolated. Incubation of this Mr 43,000 protein with isolated mitochondria resulted in processing of the Mr 43,000 precursor to the Mr 40,000 mature-sized protein. Addition of carbonyl cyanide m-chlorophenylhydrazone and/or phenanthroline inhibits this processing. These data indicate that ferrochelatase, like most mitochondrial proteins, is synthesized in the cytoplasm as a larger precursor and is then translocated and processed to a mature-sized protein in an energy-required step.

Animals↗
PubMed1983

Monoclonal antibodies as probes for differentiating tropoelastin a and b.

BALB/C mice were immunized with a mixture of aortic tropoelastin a and b. To ensure the presence of tropoelastin a, the lathyritic chick aortic tissue was first extracted with denaturing and reducing agents. After dialysis, the standard procedure for tropoelastin purification was followed. Following immunization, the lymphocytes were fused with a myeloma cell line. Resulting hybridomas were cloned and tested for reactivity towards tropoelastin a and b. The majority of monoclonal antibodies exhibited crossreactivity between the two tropoelastins. Two of the clones were further characterized because of their ability to differentiate both tropoelastin a and b. As demonstrated by immunoprecipitation and Western blot analyses, one clone reacted preferentially with tropoelastin b and the other with tropoelastin a and b. Significantly, these results demonstrate that tropoelastin a and b can be differentiated by immunological techniques.

Animals↗
PubMed1981

Primary structure of the signal peptide of tropoelastin b.

Elastin is a major protein of compliant connective tissue and is characterized by an amino acid composition abundant in nonpolar residues. The soluble precursor to elastin tropoelastin, is extractable in organic solvents and possesses an extensive clustering of nonpolar amino acid residues in the immediate NH2-terminal region (Foster, J. A., Shapiro, R., Voynow, P., Crombie, G., Faris, B., and Franzblau, C. (1975) Biochemistry 14, 857-864). It was, therefore, of special interest to determine whether tropoelastin requires a hydrophobic signal peptide for vectorial transport of the nascent polypeptide. The possibility that the initial tropoelastin translation product possesses a short signal peptide was examined in a cell-free translation system. Total RNA, isolated from aortae of 1-day-old chicks, was translated in an mRNA-dependent reticulocyte lysate translation assay. The translation products were then immunoprecipitated and subjected to automated radiosequencing. Comparison of the NH2-terminal sequence of tropoelastin b synthesized in the cell-free system versus that synthesized in organ culture demonstrated the presence of a signal peptide 24 amino acid residues in length. The signal peptide sequence is as follows: Met-Arg-Gln-Ala-Ala-Ala-Pro-Leu-Leu-Pro-Gly-Val-Leu-Leu-Leu-Phe-Ser-Ile-Leu-Pro -Ala-Ser-Gln-Gln. The preponderance of hydrophobic amino acid residues as well as the polar residues adjacent to the initiator methionine and the carboxyl termini found in the signal peptide is similar to that reported for other secreted proteins.

Amino Acid Sequence↗