Identification of sugarcane genes involved in the purine synthesis pathway

Abstract

Nucleotide synthesis is of central importance to all cells. In most organisms, the purine nucleotides are synthesized de novo from non-nucleotide precursors such as amino acids, ammonia and carbon dioxide. An understanding of the enzymes involved in sugarcane purine synthesis opens the possibility of using these enzymes as targets for chemicals which may be effective in combating phytopathogen. Such an approach has already been applied to several parasites and types of cancer. The strategy described in this paper was applied to identify sugarcane clusters for each step of the de novo purine synthesis pathway. Representative sequences of this pathway were chosen from the National Center for Biotechnology Information (NCBI) database and used to search the translated sugarcane expressed sequence tag (SUCEST) database using the available basic local alignment search tool (BLAST) facility. Retrieved clusters were further tested for the statistical significance of the alignment by an implementation (PRSS3) of the Monte Carlo shuffling algorithm calibrated using known protein sequences of divergent taxa along the phylogenetic tree. The sequences were compared to each other and to the sugarcane clusters selected using BLAST analysis, with the resulting table of p-values indicating the degree of divergence of each enzyme within different taxa and in relation to the sugarcane clusters. The results obtained by this strategy allowed us to identify the sugarcane proteins participating in the purine synthesis pathway.

Sugarcane Phosphoribosyl Pyrophosphate Synthetase: Molecular Characterization of a Phosphate-independent PRS

“…Nucleotide Sequence Analysis Previous data-mining work (Jancso et al 2001) identified seven full-length and nine partial prs clones distributed in 16 tissue-specific and one normalized sugarcane cDNA library. One of these clones (SCACRZ3035C11) from the plat root library was selected and a 1,194-bp DNA fragment was PCR amplified.…”

“…The SUCEST (http://sucest.lad.dcc.unicamp.br/en/) databank identified 16 PRS coding EST clones, seven full length and nine partial sequences, distributed in 6 of the 16 tissues that had libraries generated (Jancso et al 2001). The analysis done by Jancso et al (2001) was unable to identify a total number of PRS isoforms present in the sugarcane genome, but it is clear that a number of isoforms are present.…”

“…The analysis done by Jancso et al (2001) was unable to identify a total number of PRS isoforms present in the sugarcane genome, but it is clear that a number of isoforms are present. Probably at least three forms are expected to be present, PRS-I, PRS-II, and PRS-III, participating as a multienzymatic complex.…”

Preliminary crystallographic analysis of sugar cane phosphoribosylpyrophosphate synthase

“…The SUCEST strategy was based on the random sequencing of EST (expressed sequence tag) fragments obtained from different plant tissues and allows investigation of the expressed genes of a complex genome. Based on the knowledge generated from the SUCEST project, several important genes have been identi®ed, including representatives of the purine-salvage and de novo synthesis pathways (Jancso et al, 2001). In the present report, we describe the puri®cation, crystallization and preliminary crystallographic analysis of the sugar cane PRPP synthase enzyme as part of a continuing study of de novo and salvage pathway enzymes in several organisms.…”