title: Current functional metagenomic approaches only expand the existing protease sequence space, but does not presently add any novelty to it.
abstract: Proteases are a fundamental function in many organisms and thus many ecosystems and yet they are rarely obtained in functional metagenomic screens. Here, we have isolated an active protease gene (M1-2; 613 amino acids) which resided in a 38.4 kb fosmid clone that showed a classical protease-positive phenotype. It was classified as a zinc-dependent metalloprotease, with the closest annotated sequence as a neutral protease from Collimonas fungivorans (62 % similarity and 72 % homology). Further characterisation showed that its optimum temperature and pH were 42 °C and 8.0, respectively. Activity was inhibited by EDTA, but inhibition started to be reversed by excess Zn(2+). A putative signal peptide was identified bioinformatically and this may be why this protease was successfully isolated using a functional metagenomic screen. Bioinformatic analysis shows that this does not represent a novel protease, but simply expands the current sequence space of known proteases.
authors.affiliation: School of Biosciences, Cardiff University, Museum Avenue, Cardiff, CF10 3AX, UK.
authors.forename: Laura S
authors.initials: LS
authors.lastname: Morris
authors.forename: Julian R
authors.initials: JR
authors.lastname: Marchesi
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D056186:Metagenomics; D008969:Molecular Sequence Data; D042802:Oxalobacteraceae / Q000737:chemistry / Q000201:enzymology* / Q000235:genetics; D010447:Peptide Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D010802:Phylogeny; D017386:Sequence Homology, Amino Acid
chemical_list: D001426:Bacterial Proteins; D010447:Peptide Hydrolases