Abstract Cloning and characterisation of the cutinase genomic and cDNA gene from the fungal phytopathogen Glomerella cingulata

¹School of BioSciences and Biotechnology, Faculty of Science and Technology,
Universiti Kebangsaan Malaysia, Bangi 43600, Selangor, Malaysia;
²Institute of Molecular Biosciences, Massey University, Palmerston North, New Zealand

Received 19 November 2001 / Accepted 12 December 2001

Abstract.
The cutinase gene (cutA) has been identified from a genomic DNA library of the plant pathogenic fungus Glomerella cingulata ICMP 11061. Nucleotide sequence data revealed that this gene codes for a putative 224-amino acid protein encoded by two exons of 189 bp and 486 bp, separated by an intron of 52 bp. The presence of the 52 bp intron was confirmed after comparison with the cutinase cDNA sequence obtained using RT PCR of cutin induced cells. The DNA segments from positions-120 to-112 and from positions-101 to-95 relative to the start codon are potential segments for cutinase transcription start and transcription factor binding sites, respectively. The presumptive TATA box is mapped at-116 from the initiation of translation site. Potential adenylation sites are mapped at segments 228 to 231 and 257 to 259 downstream from the stop signal. The cutinase gene is present in a single copy in the genome of G. cingulata and the putative protein product is between 24 and 99 per cent identical at the amino acid level to other fungal cutinases. The three dimensional protein structure of G. cingulata cutinase as determined by protein homology modeling showed that the protein is ellipsoidal and has a central ß sheet consisting of five parallel strands surrounded by 5 helices. The amino acid residues potentially participating in the catalytic triad and oxyanion hole have been determined and are located at one extremity of the protein.

Keywords: Cutinase gene, Glomerella cingulata