The dipeptidyl peptidase IV family in cancer and cell biology.

Of the 600+ known proteases identified to date in mammals, a significant percentage is involved or implicated in pathogenic and cancer processes. The dipeptidyl peptidase IV (DPIV) gene family, comprising four enzyme members [DPIV (EC 3.4.14.5), fibroblast activation protein, DP8 and DP9] and two nonenzyme members [DP6 (DPL1) and DP10 (DPL2)], are interesting in this regard because of their multiple diverse functions, varying patterns of distribution/localization and subtle, but significant, differences in structure/substrate recognition. In addition, their engagement in cell biological processes involves both enzymatic and nonenzymatic capabilities. This article examines, in detail, our current understanding of the biological involvement of this unique enzyme family and their overall potential as therapeutic targets.

Functional characterization and specific effects of various peptides on enzymatic activity of a DPP-III homologue from goat brain.

The purified dipeptidyl aminopeptidase from goat brain showed several characteristics similar to DPP-III although it possesses a dissimilar molecular weight and different inhibition behavior. The enzyme was found to be inhibited by metallochelators and thiol inhibitors which could be reversed by introducing metals and thiols, respectively. The enzyme activity is also significantly affected by DMSO and ethanol. It was found to be highly sensitive to even very low concentration of urea. The inhibitory potency of several dipeptides and bioactive peptides on this enzyme was investigated to characterize its active site. The highest potency was observed for the dipeptides having aromatic and bulky side chains such as Phe-Met, Leu-Arg, Met-Arg, Trp-Met and Leu-Trp.

Markers for host-induced gene expression in Trichophyton dermatophytosis.

Dermatophytes are adapted to infect keratinized tissues by their ability to utilize keratin as a nutrient source. Although there have been numerous reports that dermatophytes like Trichophyton sp. secrete proteolytic enzymes, virtually nothing is known about the patterns of gene expression in the host or even when the organisms are cultured on protein substrates in the absence of a host. We characterized the expression of an aminopeptidase gene, the Trichophyton mentagrophytes homolog of the Trichophyton rubrum Tri r 4 gene. The T. rubrum gene was originally isolated based on the ability of the protein encoded by it to induce immediate and delayed-type hypersensitivity in skin tests. T. mentagrophytes Tri m 4 is closely related to Tri r 4 (almost 94% identity at the protein level). Tri m 4 resembles other protease-encoding genes thought to be virulence factors (for example, DPP V of Aspergillus fumigatus). The Tri m 4 protein was detected immunochemically both in fungal extracts and in the culture medium. Expression of the Tri m 4 gene was induced severalfold when T. mentagrophytes was grown on keratin and elastin. Ex vivo, strong induction was observed after culture on blood plasma, but the use of homogenized skin did not result in a significant increase in Tri m 4 transcript levels. In order to identify additional genes encoding putative virulence factors, differential cDNA screening was performed. By this method, a fungal thioredoxin and a cellulase homolog were identified, and both genes were found to be strongly induced by skin extracellular matrix proteins. Induction by superficial (keratin) and deep (elastin) skin elements suggests that the products of these genes may be important in both superficial and deep dermatophytosis, and models for their function are proposed. Upregulation of several newly identified T. mentagrophytes genes on protein substrates suggests that these genes encode proteins which are relevant to the dermatophyte-skin interaction.

Characterisation of the Aspergillus niger dapB gene, which encodes a novel fungal type IV dipeptidyl aminopeptidase.

We have cloned the Aspergillus niger dapB gene. Analysis of its nucleotide sequence and the corresponding protein sequence indicates that the gene encodes a type IV dipeptidyl aminopeptidase (DPP IV). Based upon its deduced sequence we predict the presence of a transmembrane domain in the protein. Furthermore, dapB-overexpressing transformants display an increase in intracellular DPP IV activity. This is the first reported characterisation of a dipeptidyl aminopeptidase with a transmembrane domain from a filamentous fungus. Using the dapB sequence as a query, we were able to identify 14 DPP IV-encoding genes, and 12 additional DPPIV proteases in public genomic databases. Phylogenetic analysis reveals that in yeasts there are two clades of genes that encode DPP IV proteases with a transmembrane domain. In this study we demonstrate that, as in yeasts, two classes of DPP IV-encoding genes exist in filamentous fungi. However, only one of these codes for DPP IV proteases with a transmembrane domain. The second type present in filamentous fungi encodes extracellular DPP IV proteases. The dapB gene belongs to the first cluster. We propose that DapB plays a role in the proteolytic maturation of enzymes produced by A. niger.

Dipeptidyl peptidase with strict substrate specificity of an anaerobic periodontopathogen Porphyromonas gingivalis.

A dipeptidyl peptidase which hydrolyzed Xaa-Ala-p-nitroanilide was purified to homogeneity by sequential procedures including ammonium sulfate precipitation, ion-exchange chromatography, hydrophobic interaction chromatography, gel filtration and isoelectric focusing from the cell extract of Porphyromonas gingivalis. The purified enzyme hydrolyzed p-nitroanilide derivatives of Lys-Ala, Ala-Ala, and Val-Ala, but not Xaa-Pro. Enzyme activity was maximum at neutral pHs. Its molecular mass was 64 kDa with an isoelectric point of 5.7. The enzyme belonged to the family of serine peptidases.