Molecular characterization of an olive imidazoleglycerol-phosphate dehydratase
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Imidazoleglycerol-phosphate dehydratase (IGPD) catalyzes the6th step in histidine biosynthesis in various organism includingfungi, bacteria and plants. There are two different isoforms ofIGPD; IGPD1 and IGPD2. By finding specific inhibitors ofIGPD in plants, many herbicides have been designed. Althoughthere are numerous reports on various plantIGPDs, there are nostudies reporting olive IGPD. The aim of this study is to conductcharacterization of an oliveIGPD(OeIGPD).OeIGPDcDNA was isolated from a cDNA library we con-structed from olive pedicels. Homology searches for nucleotide,amino acids and alternative open reading frames were conductedutilizing BLASTn, BLASTp, and BLASTx, respectively. Nucleo-tide sequences of homologous genes from other plants werealigned using BioEdit and the number of SNPs were detected.The alignment was then used to generate a phylogenetic tree usingMEGA7 program. Another alignment with amino acid sequencesof the homologues proteins was also generated to construct a phy-logenetic tree displaying OeIGPD’s position among other plants.Various aspects of OeIGPD including amino acid composi-tion, hydropathy analysis, isoelectric point (pI) and three dimen-tional structure of the protein were determined using online software at ExPASY. Multiple primer pairs to amplify the fulllength open reading frame of the gene, to clone the gene into theexpressing vector pLATE51, and to detect expression throughreal-time PCR were designed using Primer3. Amino acid compo-sition analysis revealed that OeIGPD contained serine, arginineand isoleucine predominantly while hydropathy analysis sug-gested it was an hydrophilic protein. Isoelectric point (pI) of theprotein was calculated as 4.97. The molecular weight of the pro-tein was calculated as 11 kDa. Analyses continue to determinethe polymorphism ofOeIGPDamong olive cultivars, and bio-chemical function of the gene in olive.