A Novel β-Peptidyl Aminopeptidase (BapA) from Strain 3-2W4 Cleaves Peptide Bonds of Synthetic β-Tri- and β-Dipeptides

Abstract

A novel bacterial strain that was capable of growing on the ␤-tripeptide H-␤hVal-␤hAla-␤hLeu-OH as the sole carbon and nitrogen source was isolated from an enrichment culture. On the basis of physiological characterization, partial 16S rRNA sequencing, and fatty acid analysis, strain 3-2W4 was identified as a member of the family Sphingomonadaceae. Growth on the ␤-tripeptide and the ␤-dipeptide H-␤hAla-␤hLeu-OH was observed, and emerging metabolites were characterized. Small amounts of a persisting metabolite, the N-acetylated ␤-dipeptide, were identified in both media. According to dissolved organic carbon measurements, 74 to 80% of the available carbon was dissimilated. The ␤-peptide-degrading enzyme was purified from the crude cell extract of cells from strain 3-2W4 grown on complex medium. The enzyme was composed of two subunits, and the N-terminal sequences of both were determined. With this information, it was possible to identify the complete nucleotide sequence and to deduce the primary structure of the gene bapA. The gene encoded a ␤-peptidyl aminopeptidase (BapA) of 402 amino acids that was synthesized as preprotein with a signal sequence of 29 amino acids. The enzyme was cleaved into two subunits (residues 30 to 278 and 279 to 402). It belonged to the N-terminal nucleophile (Ntn) hydrolase superfamily.

Description of Sphingosinicella xenopeptidilytica sp. nov., a β-peptide-degrading species, and emended descriptions of the genus Sphingosinicella and the species Sphingosinicella microcystinivorans

“…Strain 3-2W4 T and Sphingosinicella microcystinivorans Y2 T utilized many amino acids and some organic acids as sources of carbon and energy, but they did not grow on any of the sugars and polyalcohols tested ( Table 2). The only characteristics that distinguish strain 3-2W4 T from Sphingosinicella microcystinivorans Y2 T concerned the intensity of growth on b-homovaline, citrate and DL-lactate and the reduction of nitrate to nitrite (Table 2).Recently, the b-peptidyl aminopeptidase of strain 3-2W4 T was isolated and characterized and its coding sequence (bapA) analysed (Geueke et al, 2005). This enzyme is responsible for the ability of strain 3-2W4 T to utilize the synthetic b-peptides H-bhVal-bhLeu-bhAla-OH and HbhAla-bhLeu-OH as sole sources of carbon, nitrogen and energy (Geueke et al, 2005).…”

“…Recently, the b-peptidyl aminopeptidase of strain 3-2W4 T was isolated and characterized and its coding sequence (bapA) analysed (Geueke et al, 2005). This enzyme is responsible for the ability of strain 3-2W4 T to utilize the synthetic b-peptides H-bhVal-bhLeu-bhAla-OH and HbhAla-bhLeu-OH as sole sources of carbon, nitrogen and energy (Geueke et al, 2005).…”

“…Although b-peptides are of synthetic origin and exhibit extremely high resistance to enzymic degradation, different bacterial consortia have been shown to have the capacity to grow on these substrates (Schreiber et al, 2002). Recently, strain 3-2W4 T was isolated from one of these mixed cultures and it was shown that this strain utilized the b-tripeptide H-bhVal-bhAla-bhLeu-OH as a sole source of carbon and energy (Geueke et al, 2005). The inoculum for the enrichment culture originated from the aeration tank of a wastewater treatment plant at Leutschenbach in Zurich, Switzerland.…”

“…This enzyme is responsible for the ability of strain 3-2W4 T to utilize the synthetic b-peptides H-bhVal-bhLeu-bhAla-OH and HbhAla-bhLeu-OH as sole sources of carbon, nitrogen and energy (Geueke et al, 2005). The two b-peptides were synthesized according to Schreiber et al (2002).…”

Biodegradable Polymers: Recent Developments and New Perspectives

“…PahZ1 enzymes are classified into the α/β-hydrolase_5 family, which includes putative PHB depolymerase (LpqC) from Bordetella parapertussis, based on the ESTHER database (http://bioweb.ensam.inra.fr/esther). As shown in Figure 10, the residues composing their catalytic triads, in which the Ser residue formed together with Asp and His residues, are conserved in PahZ1 KT In addition to the PAA-hydrolysing enzymes (PahZ1 KT-1 , PahZ2 KT-1 , and PahZ1 KP-2 ), five β-aminopeptidases (three BapA enzymes, one BapF enzyme, and one DmpA enzyme) hydrolyse short β-peptides and β-amino-acid containing peptides [135][136][137][138][139][140][141][142]. Their properties are listed in Table 2.…”