Identification of a new phospholipase D in Carica papaya latex

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• 作者：Slim Abdelkafi^a ; ^b ; ^{slim.abdelkafi@enis.rnu.tn} ; Abdelkarim Abousalham^c ; Imen Fendri^b ; Hiroyuki Ogata^d ; Nathalie Barouh^e ; Benjamin Fouquet^f ; Frantz Scheirlinckx^f ; Pierre Villeneuve^e ; Fré ; dé ; ric Carriè ; re^a
• 关键词：CpPLD1 ; Carica papaya phospholipase ; D1 ; phospholipase D ; EST ; expressed sequence tag ; EtOH ; ethanol ; MS ; mass spectrometry ; ORF ; open reading frame ; PA ; phosphatidic acid ; PEtOH ; phosphatidylethanol ; PLD ; PtdCho ; phosphatidylcholine ; SDS-PAGE ; sodium dod
• 刊名：Gene
• 出版年：2012
• 期刊代码：73_03781119
• 类别：bio
• 出版时间：15 May, 2012
• 卷：499
• 期：2
• 页码：243-249
• 文件大小：727 K

摘要

Phospholipase D (PLD) is a lipolytic enzyme involved in signal transduction, vesicle trafficking and membrane metabolism. It catalyzes the hydrolysis and transphosphatidylation of glycerophospholipids at the terminal phosphodiester bond. The presence of a PLD in the latex of Carica papaya (CpPLD1) was demonstrated by transphosphatidylation of phosphatidylcholine (PtdCho) in the presence of 2%ethanol. Although the protein could not be purified to homogeneity due to its presence in high molecular mass aggregates, a protein band was separated by SDS-PAGE after SDS/chloroform-methanol/TCA-acetone extraction of the latex insoluble fraction. This material was digested with trypsin and the amino acid sequences of the tryptic peptides were determined by micro-LC/ESI/MS/MS. These sequences were used to identify a partial cDNA (723 bp) from expressed sequence tags (ESTs) of C. papaya. Based upon EST sequences, a full-length gene was identified in the genome of C. papaya, with an open reading frame of 2424 bp encoding a protein of 808 amino acid residues, with a theoretical molecular mass of 92.05 kDa. From sequence analysis, CpPLD1 was identified as a PLD belonging to the plant phosphatidylcholine phosphatidohydrolase family.