The SCO2299 gene from Streptomyces coelicolor A3(2) encodes a bifunctional enzyme consisting of an RNase H domain and an acid phosphatase domain

Abstract

The SCO2299gene from Streptomyces coelicolor encodes a single peptide consisting of 497 amino acid residues. Its N‐terminal region shows high amino acid sequence similarity to RNase HI, whereas its C‐terminal region bears similarity to the CobC protein, which is involved in the synthesis of cobalamin. The SCO2299 gene suppressed a temperature‐sensitive growth defect of an Escherichia coli RNase H‐deficient strain, and the recombinant SCO2299 protein cleaved an RNA strand of RNA·DNA hybrid in vitro. The N‐terminal domain of the SCO2299 protein, when overproduced independently, exhibited RNase H activity at a similar level to the full length protein. On the other hand, the C‐terminal domain showed no CobC‐like activity but an acid phosphatase activity. The full length protein also exhibited acid phosphatase activity at almost the same level as the C‐terminal domain alone. These results indicate that RNase H and acid phosphatase activities of the full length SCO2299 protein depend on its N‐terminal and C‐terminal domains, respectively. The physiological functions of the SCO2299 gene and the relation between RNase H and acid phosphatase remain to be determined. However, the bifunctional enzyme examined here is a novel style in the Type 1 RNase H family. Additionally, S. coelicolor is the first example of an organism whose genome contains three active RNase H genes.

Biochemical Characterization of Mycobacterium smegmatis RnhC (MSMEG_4305), a Bifunctional Enzyme Composed of Autonomous N-Terminal Type I RNase H and C-Terminal Acid Phosphatase Domains

“…They are characterized by a signature active-site peptide motif, present in RnhC as 168 FLLRHGQT 176 , in which His173 is the putative histidine nucleophile. Recombinant full-length wild-type RnhC readily hydrolyzed p-nitrophenylphosphate to p-nitrophenol at pH 5.5 (the optimum pH reported for the S. coelicolor homolog [20]) without a divalent cation cofactor (Fig. 8A), with an apparent turnover of 4.8 min Ϫ1 .…”

“…1A). It is homologous to Streptomyces coelicolor SCO2299, a bifunctional nuclease-phosphatase enzyme with bona fide RNase H activity in vitro that is inherent to an autonomous N-terminal domain, which is as active in this regard as full-length SCO2299 (20). The C-terminal domain of SCO2299 has a generic phosphatase activity that hydrolyzes p-nitrophenyl phosphate (20).…”

“…It is homologous to Streptomyces coelicolor SCO2299, a bifunctional nuclease-phosphatase enzyme with bona fide RNase H activity in vitro that is inherent to an autonomous N-terminal domain, which is as active in this regard as full-length SCO2299 (20). The C-terminal domain of SCO2299 has a generic phosphatase activity that hydrolyzes p-nitrophenyl phosphate (20). The M. tuberculosis homolog Rv2228c was characterized by Watkins and Baker (14), who reported that (i) the full-length Rv2228c was equally adept at cleaving RNA:DNA and RNA:RNA duplexes and (ii) the isolated N-terminal RNase H domain was 100-fold less active than the full-length Rv2228c.…”

MS_RHII-RSD, a Dual-Function RNase HII-(p)ppGpp Synthetase from Mycobacterium smegmatis

“…There are instances in the literature of an RNase H domain fused to a functionally distinct non-RNase H domain, like the RNase HI-CobC acid phosphatase fusion proteins (38,57). Protein BLAST search shows that the M. smegmatis genome encodes an RNase HI-CobC fusion protein (MSMEG_4305).…”

Structural and Functional Characterization of an RNase HI Domain from the Bifunctional Protein Rv2228c from Mycobacterium tuberculosis

“…It has 34% sequence identity with the ␣-ribazole phosphatase CobC of Synechococcus sp., but it is also homologous with PhoE from Bacillus subtilis (34% identity) and Rv3214 from M. tuberculosis (28% identity), both of which have acid phosphatase activity (39,46). Bifunctional proteins similar to Rv2228c are encoded by the genomes of other Actinomycetales bacteria, including those of the Mycobacterium, Streptomyces, Corynebacterium, and Nocardia genera, and one of these bifunctional proteins, SCO2299 from Streptomyces coelicolor, has RNase HI activity in its N-terminal domain and acid phosphatase activity in its C-terminal domain (34).…”