Activation of the urease of SCHIZOSACCHAROMYCES POMBE by the UreF accessory protein from soybean

Abstract

Plant orthologs of the bacterial urease accessory genes ureD and ureF, which are required for the insertion of the nickel ion at the active site, have been isolated from soybean ( Glycine max L. Merr.), tomato ( Lycopersicon esculentum) and Arabidopsis thaliana. The functionality of soybean UreD and UreF was tested by measuring their ability to complement urease-negative mutants of Schizosaccharomyces pombe, a eukaryote which produces a "plant-like" urease of ~90 kDa. The S. pombe ure4 mutant was complemented by a 12-kb fragment of S. pombe genomic DNA, which was shown by PCR to contain a putative ureD gene. However, ure4 was not complemented by a UreD cDNA soybean, expressed under the control of a strong promoter. In contrast, an S. pombe ure3 mutation was complemented by both a 10-kb fragment of S. pombe DNA containing ureF and the UreF cDNA from soybean. Soybean Eu2 is a candidate urease accessory gene; its product cooperates with the Eu3 protein in activating apourease in vitro. However, the sequences of UreD and UreF transcripts from two eu2/eu2 mutants, recovered as RT-PCR products, revealed no mutational alteration, suggesting that Eu2 encodes neither UreD nor UreF.

Identification of Three Urease Accessory Proteins That Are Required for Urease Activation in Arabidopsis

“…In contrast, potato (Solanum tuberosum), tomato (Lycopersicon esculentum), several other solanaceous species, and also Arabidopsis (Arabidopsis thaliana; ecotype Columbia [Col]) possess only a single urease gene (Witte et al, 2005). Using microbial accessory protein sequences, putative plant orthologs for UreD, UreF, and UreG were identified in the databases and the corresponding cDNAs cloned (Freyermuth et al, 2000;Bacanamwo et al, 2002). Sequence identities determined from pair-wise alignments of K. aerogenes UreD and UreF with Arabidopsis UreD and UreF candidates are about 20% in both cases, providing only a weak indication that these proteins may be orthologous.…”

“…Functional complementation of defective microbial urease activation complexes indicated a possible role of other putative plant accessory protein orthologs in urease activation. A K. aerogenes urease operon carrying a mutation in ureG could be partially complemented by a potato ureG ortholog ) and a Schizosaccharomyces pombe ureF mutant was partially complemented by a soybean protein with similarity to microbial UreF (Bacanamwo et al, 2002). However, a role in plant urease activation still needs to be established for possible plant UreF orthologs.…”

“…However, a role in plant urease activation still needs to be established for possible plant UreF orthologs. Attempts to complement ureD mutants of K. aerogenes or S. pombe with putative plant UreD orthologs have failed so far (Witte, 2001;Bacanamwo et al, 2002). A further mutant lacking all urease activities in soybean is encoded by the Eu2 gene .…”

Identification of a novel gene, URE2, that functionally complements a urease-negative clinical strain of Cryptococcus neoformans

“…Urease accessory proteins, such as UreD, UreE and UreF, have been reported in Lycopersicon esculentum (tomato), Glycine max (soybean), Arabidopsis thaliana (thale cress) and the fission yeast Schizosaccharomyces pombe (Bacanamwo et al, 2002). Although the DNA sequences of the UreD and UreF genes appear to be well conserved among plants and some species of divergent bacteria, the homology of these genes appears to be intermediate when compared to the fungus S. pombe (Bacanamwo et al, 2002). The accessory proteins UreD and UreF are apparently essential for urease activity and reportedly form a complex to which the chaperone protein UreE delivers nickel, thus activating the urease enzyme (Lee et al, 1992;Soriano et al, 2000).…”

“…The accessory proteins UreD and UreF are apparently essential for urease activity and reportedly form a complex to which the chaperone protein UreE delivers nickel, thus activating the urease enzyme (Lee et al, 1992;Soriano et al, 2000). Synthesis of metalcontaining enzymes, such as urease, which requires the binding of nickel for function (Bacanamwo et al, 2002;Eitinger et al, 2000), often elicits the participation of several accessory proteins. It is postulated that the UreD gene is in some way involved in the production or incorporation of a nickel co-factor essential for urease activity.…”