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M Poonian

Publications and source records attributed to M Poonian.

7 recordsLinked to original sources
PubMed1985

Olfactory neuron-specific protein is translated from a large poly(A)+ mRNA.

Poly(A)+ mRNA was isolated from rat olfactory mucosa and translated in a rabbit reticulocyte cell-free protein synthesizing system. Olfactory marker protein (OMP) of Mr 18,500 was faithfully produced by this system upon addition of mucosal mRNA. The protein was identified by radioimmunoprecipitation with specific anti-OMP serum and by competitive displacement of the radioactive product with authentic OMP. In addition, the immunoprecipitated product comigrated with OMP on NaDodSO4/polyacrylamide gels and on HPLC. In vitro synthesized OMP represented 0.5% of the total translational products. Total olfactory mucosal poly(A)+ mRNA is approximately 1.5-21 kilobases in size, as determined by denaturing agarose gels. Translational assays of gel-fractionated poly(A)+ mRNA demonstrated that OMP mRNA occurs in the 2.5- to 3.4-kilobase range. An mRNA of this size could code for a protein significantly larger than OMP. Since the in vitro synthesized OMP is indistinguishable in size from OMP isolated from tissue, our data indicate that OMP is synthesized directly without the intermediate formation of a larger polypeptide precursor. Thus, OMP mRNA contains untranslated regions that are four to five times larger than the coding region.

Animals↗
PubMed1985

Molecular cloning and regulated expression of the human c-myc gene in Escherichia coli and Saccharomyces cerevisiae: comparison of the protein products.

mRNA from human HL-60 cells was used to prepare a cDNA library, from which two full-length clones that encompass the complete c-myc coding region were isolated. One clone, pM1-11, contains all three exons of human c-myc. The second clone, pM4-10, represents a relatively rare transcript that initiated in the first intron and includes the coding exons 2 and 3. The cDNA insert in pM1-11 was used to express the human c-myc protein in both prokaryotic and eukaryotic cells. Insertion of the coding sequences in exons 2 and 3 into the appropriate expression vectors yielded detectable c-myc protein in Escherichia coli lacking the Lon protease and in Saccharomyces cerevisiae upon induction. The protein produced in E. coli has an apparent size of 60 kDa and appears to be unmodified, as it is identical in size to the protein synthesized in an in vitro system. In contrast, yeast cells synthesize two myc proteins, of 60 kDa and 62 kDa. The difference in apparent molecular mass between the two proteins appears to be due, in part, to phosphorylation. Subcellular fractionation of yeast cells showed that the c-myc phosphoprotein is located predominantly in the nuclear fraction.

Base Sequence↗
PubMed1984

Molecular cloning and sequence determination of rat preproenkephalin cDNA: sensitive probe for studying transcriptional changes in rat tissues.

A cDNA probe was prepared to investigate the regulation of proenkephalin biosynthesis in the rat. This was necessary because human and bovine proenkephalin cDNA were not sensitive enough for the accurate detection of preproenkephalin mRNA in tissues that contain low copy numbers of this message, such as the adrenal gland. The rat probe was prepared in the following manner. Preproenkephalin mRNA was enriched by sucrose gradient centrifugation of poly(A)-containing mRNA from rat brain and was used as a template for double-stranded cDNA synthesis. The resulting cDNA was inserted into the plasmid pBR322, and recombinant plasmids were used to transform Escherichia coli RR1 cells. A synthetic oligodeoxyribonucleotide (30 bases long) with a sequence that had previously been shown to be identical in bovine and human preproenkephalin cDNA was prepared to screen the clone bank. The plasmid with the longest cDNA insert (about 1200 bases) from the positive clones was isolated, and the sequence of the entire protein coding region was determined. Like the bovine and human gene products, rat preproenkephalin contains four [Met]enkephalin sequences and one copy each of [Leu]enkephalin, [Met]enkephalin-Arg6-Gly7-Leu8, and [Met]enkephalin-Arg6-Phe7. Rat preproenkephalin is 80% and 83% homologous to the bovine and human forms, respectively, at the nucleotide level and is 82% homologous to both species at the amino acid level. Rat preproenkephalin contains 269 amino acid residues, making it larger than the human (267 residues) and bovine (263 residues) precursors. The sensitivity for detection of rat preproenkephalin mRNA with the rat cDNA was several times greater than with the corresponding cDNAs from bovine and human sources.

Amino Acid Sequence↗