Isolation of one novel full-length gene PPIL-3 related to human glioma and characterization of this protein

Abstract

Objective To obtain differentially expressed genes related to human glioma using cDNA microarray and the characterization of one novel full-length gene. Method Total RNA was extracted from human glioma tissues and normal brain tissues,and mRNA was used to make probes. After hybridization and washing procedure, the results of hybridization were scanned using computer system. One gene named 681F05 clone was subsequently analyzed by northern blot, four times hybridizations and scanning. Northern blot analysis confirmed 681F05 clone was low-exprbioinformatics and protein expression. Results We obtain 15 differentially expressed genes to human glioma througession in human brain tissue and over-expression inhuman glioma tissues. The analysis of BLASTn and BLASTx showed that clone 681F05 we isolated was two cDNA clones encoding two novel proteins which show 52% and 72% identity to the eyelophilin isoform 10 of C. Elgans,respectively. Sequence analysis revealed these two cDNA clones are two different splicing variants of a novel cycophilin-like gene (PPIL3a and PPIL3b). In E. Coli. ,we got the more high-expressed fusion protein of PPIL3a combined with GST and PPIL3b combined with GST. Activity analysis of the recombinant PPIL3b protein shows that PPIL3b protein has nuclease activity. The nuclease activity of PPIL3b protein is stimulated by Ca2+ and/or Mg2+ and inhibited by K+/Na+. Conclusions We show that cDNA microarray technology can be successfully applied to identify differentially expressed genes. The novel full-length gene of human PPIL3 maybe correlate with forming human glioma. Key words: cDNA microarray; Glioma; PPIL-3; Gene