Organic compounds -- part of the class 532-570 series – Organic compounds – Carbohydrates or derivatives
Patent
1997-10-17
1999-11-16
Achutamurthy, Ponnathapura
Organic compounds -- part of the class 532-570 series
Organic compounds
Carbohydrates or derivatives
435195, 4352523, 4353201, 530350, C07N 2104, C12N 914, C12N 120, C12N 1500
Patent
active
059860804
ABSTRACT:
We have cloned and sequenced the cDNA encoding the 61 kD active fragment of a unique porcine chondrocyte nucleotide pyrophosphohydrolase (NTPPHase) from a porcine chondrocyte library. Degenerate oligonucleotides, corresponding to the N-terminal amino acid sequence of this peptide were hybridized to porcine chondrocyte cDNA and used to amplify DNA encoding the N-terminal sequence of 61 kD with the polymerase chain reaction (PCR). The PCR products were then used as probes to clone the entire open reading-frame for the 61 kD fragment from a porcine chondrocyte cDNA library. The length of the cloned cDNA was 2509 bp. Translation of the open-reading-frame predicts the 61 kD fragment to be a 459 amino acid protein. BLAST and FASTA analysis confirmed that this amino acid sequence was unique and did not possess high homology to any known proteins in the non-redundant protein data base. Limited homology (17%) between the 61 kD fragment and several prokaryotic and eukaryotic ATP pyrophosphate-lyase (adenylate cyclase) was detected. Northern blot analysis of porcine chondrocyte RNA showed that the DNA encoding the 61 kD fragment hybridized to a 4.3 kbp RNA transcript. Human chondrocyte RNA also hybridized to this porcine DNA probe. Coupled in vitro transcription translation of an expression vector containing the DNA insert in frame showed the expression of a 61 kD protein.
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Barbieri Joseph T.
Haas Arthur L.
Halligan Brian D.
Masuda Ikuko
McCarty Daniel J.
Achutamurthy Ponnathapura
MCW Research Foundation
Saidha Tekchand
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