We have cloned and characterized the gene of is a single-copy gene encoding a proteins homologous to fungal serine endoproteases, which localize towards the Golgi equipment. attempts to look for the 5 end from the cDNA sequence by 5 RACE were unsuccessful and were limited by our ability to obtain only small amounts of undegraded RNA from autopsy lung samples. However, based on a comparison with the genomic sequence, the cDNA sequence appears to include the complete coding sequence. There are two potential initiation codons, at nucleotide positions 536 and 539 of the genomic sequence. Although the second one appears to be the more favorable translation start site (9), we are unable to determine which codon is in fact utilized. The stop codon TAA is at position 3245. Comparison of the cDNA with the genomic sequence identified eight introns whereas, in comparison, in this gene is reported elsewhere to have only seven introns (12, 17). The cDNA encodes a protein containing 779 amino acids. Comparison of the deduced amino acid sequence with those of other fungal serine endoproteases (Fig. ?(Fig.1)1) allowed identification PHA-767491 of characteristic domains, including a signal peptide, a prodomain, a subtilisin-like catalytic domain, a P domain, a serine/threonine-rich region, and a transmembrane domain (4, 23). A hydrophobic transmembrane domain followed by a hydrophilic intracytoplasmic region, as predicted by the transmembrane hidden Markov model (18), is present at the carboxy terminus and is characteristic of Golgi apparatus-associated yeast kexins (7). FLJ12788 This is different from most PRT-1 clones of rat-derived but is similar to clone 71 (17) as well as Kex1 from mouse-derived (11). The prodomain that PHA-767491 can be removed by autocatalytic cleavage has a potential cleavage site (KR) at amino acid positions 113 and 114 (6). The deduced amino acid sequence of Kex1 from showed 39% identity to surface protease (SPRT) clone 12; PHA-767491 37% identity to clone 71; and 34% identity to Kex1 of and that is absent in Kex1 of as well as other fungal kexins (2, 7, 12, 17). FIG. 1. Comparison of the deduced amino acid sequence of Kex1 from human-derived with those of other fungal kexin-like proteases. Sequences of Kex1 of (HPC Kex1), Kex1 of (MPC Kex1), SPRTs (clone 71 and clone 12) of … Northern blot analysis showed that mRNA is 2.4 kb in size. To determine the copy number of from from was used as the probe. Southern blotting showed a single band, demonstrating that’s present like a single-copy gene in duplicated the single-copy evidently, presumably Golgi equipment type of PRT-1 (with a straightforward frameshift to improve to a glycosylphosphatidylinositol-type anchor) (17) sooner or later following the divergence of the two strains. Actually, the duplication will need to have occurred following the divergence from the in any other case very carefully related varieties, and varieties examined to day consist of multiple copies of MSG genes, whose duplication will need to have occurred in an exceedingly early ancestor of all or all the present-day varieties (5, 8, 24). FIG. 2. Southern blot evaluation of DNA from PCR item. Molecular pounds markers are demonstrated on the remaining. In are even more sensitive compared to the crazy PHA-767491 type to antifungal medicines and chemicals focusing on the cell membrane (1). Kex2 may be mixed up in control of protein that keep up with the cell surface area integrity. Therefore, it has been proposed elsewhere that Kex2 inhibitors along with other antifungal brokers could be useful for the treatment of fungal infections (1). The role of Kex1 of in processing proteins that maintain cell surface integrity remains to be investigated. Kex1 may also be involved in the proteolytic processing of MSG, which is usually important in the pathogenicity of this organism, by removing the invariant upstream conserved sequence whose expression on the surface of the organism would counteract the antigenic diversity provided by the variant MSGs (22). Thus, inhibition of Kex1 may provide a new therapeutic approach to the management of pneumonia. Nucleotide sequence accession number. The genomic sequence of was.