A Pneumocystis carinii multi-gene family with homology to subtilisin-like serine proteases

Abstract

Medicine, John Radcliffe Hospita't Oxford OX3 9DUtCopies of a multi-gene family, named PRTl (protease 3, encoding a subtilisinlike serine protease were cloned from the opportunistic fungal pathogen Pneumocystis carinii. Comparison of the nucleotide sequence of a genomic clone and a cDNA clone of PRTl from P. carinii f. sp. carinii revealed the presence of seven short introns. Several different domains were predicted from the deduced amino acid sequence: an N-terminal hydrophobic signal sequence, a pro-domain, a subtilisin-like catalytic domain, a P-domain (essential for proteolytic activity), a proline-rich domain, a serinerthreoninerich domain and a C-terminal hydrophobic domain. The catalytic domain showed high homology to other eukaryotic subtilisin-like serine proteases and possessed the three essential residues of the catalytic active site. Karyotypic analysis showed that PRTl was a multi-gene family, copies of which were present on all but one of the P. carinii f. sp. carinii chromosomes. The different copies of the PRTl genes showed nucleotide sequence heterogeneity, the highest level of divergence being in the proline-rich domain, which varied in both length and composition. Some copies of PRTl were contiguous with genes encoding the P. carinii major surface glycoprotein. UK

Purification of a 68-kDa cysteine proteinase from crude extract of Pneumocystis carinii

“…The subtilisin-like serine proteinases of P. carinii was confirmed and its gene has been cloned (Lugli et al 1997;Wada and Nakamura, 1997). The proteinase may be involved in cleavage of major surface glycoproteins or other related proteins of P. carinii (Wada and Nakamura, 1997).…”

Colonization by Pneumocystis jirovecii and Its Role in Disease

“…Recently, a second Pneumocystis protein has been investigated for potential use in serologic studies (102,154,219). The Pneumocystis protease, kexin (Kex1, Prt1), possesses similarity to the fungal subtilisin-like serine proteases Kex1 from Kluyveromyces lactis and Kex2 from Saccharomyces cerevisiae and Schizosaccharomyces pombe (165,173,181,182). The Pneumocystis-derived kex1 gene isolated from P. murina (173), as well as that from P. jirovecii (165), is a single-copy gene, whereas a multigene kex1 family has been characterized in rat-derived P. carinii (279).…”

Characterization of variants of the gene encoding the p55 antigen in Pneumocystis from rats and mice

“…Whilst PCR or subcloning artefacts could have generated some of the variants that we have identified, identification of the same variants in organisms obtained from two separate animals strongly suggests that all variants are real. Whilst additional studies are needed to better characterize these variants, to try to identify additional variants and to localize the variants on chromosomes, it seems likely, based on variable sequence data as well as Southern and Northern blot studies, that p55 is encoded by multiple genes, similar to two other surface proteins, MSG and PRT-1 (Kovacs et al, 1993;Lugli et al, 1997;Russian et al, 1999). The presence of multiple p55 genes supports a hypothesis that was derived from studies of MSG (Angus et al, 1996;Kovacs et al, 1993), i.e.…”

“…AF494450), which contained a single incomplete ORF encoding a 269 aa protein that was homologous to the previously reported P. carinii p55 gene and protein (Smulian et al, 1992(Smulian et al, , 1993, with respectively 76 and 60 % sequence identity in overlapping regions. Given that MSG and another P. carinii surface protein, PRT-1, are encoded by multi-copy gene families (Kovacs et al, 1993;Lugli et al, 1997;Russian et al, 1999), we wanted to determine whether p55-related proteins are encoded by multiple genes. We amplified P. carinii genomic DNA from a single rat isolate by PCR with primers (PU947 and PD1637; Table 1) that were based on regions conserved between these two sequences.…”