Regulated expression of the histidase structural gene in Streptomyces griseus

Abstract

The histidase structural gene from Streptomyces griseus was expressed from a leaderless, monocistronic transcript. Multiple copies of the DNA located upstream of the hutH transcription initiation site led to a significant level of histidase activity when present in trans in the wild-type strain grown under noninducing conditions.In bacteria that utilize L-histidine as a carbon or nitrogen source, L-histidine is first deaminated by histidine ammonia lyase (histidase [20,24,26]), which is encoded by hutH. The ammonia released by histidase enters pathways for its assimilation. The other product of deamination, urocanate, is catabolized in several steps to form L-glutamate (20, 24). Urocanase, encoded by hutU, is required for the conversion of urocanate to imidazolonepropionate in this pathway (17,20,24).Examination of the regulated synthesis of histidase and urocanase by Bacillus subtilis, Klebsiella aerogenes, and Pseudomonas putida has led to the identification of one (28), two (6, 30), and three (9, 13) contiguous hut operons, respectively, that encode the enzymes required for conversion of L-histidine to L-glutamate. Although the order of hut genes differs among these species, hutH and hutU invariably lie within the same operon. Expression of these hut genes is regulated either by a repressor protein that binds at a site adjacent to the promoter and reduces initiation of transcription (12,34) or by a mechanism that appears to involve transcription antitermination (28). Expression of hut genes in these bacteria is also controlled by the global mechanisms of carbon catabolite repression (27,29,37) and nitrogen regulation (25,31) or by amino acid repression (1).Previous studies have demonstrated that L-histidine is a good nitrogen source for Streptomyces coelicolor (20) and Streptomyces griseus (22). In streptomycetes the enzymes of histidine catabolism are active whenever L-histidine or urocanate is present in the medium, regardless of the presence of other carbon or nitrogen sources (3,20,22). Thus, there is no evidence for carbon or nitrogen regulation of the histidine utilization (Hut) system in streptomycetes. On the basis of enzymological analyses, a Hut Ϫ mutant that we isolated appeared to constitutively synthesize inactive histidase that could be activated in vitro by exposure to an extract prepared from the wild-type strain (23). Earlier studies also indicated that histidase from S. griseus and S. coelicolor is a hysteretic enzyme that undergoes activation during the course of the reaction (19,22). In S. griseus, the hysteretic status of histidase appears to be dependent on the life cycle (22). Because this result suggested the possibility that the metabolic flux of L-histidine is regulated at least in part by the hysteretic activity of histidase, we wished to determine if histidase synthesis is also regulated in S. griseus.The histidase structural gene from the wild-type strain of S. griseus has been cloned, and its nucleotide sequence has been determined (38). Here we describe our analysis of hutH expr...

Substrate induction and glucose repression of maltose utilization by Streptomyces coelicolor A3(2) is controlled by malR, a member of the lacI–galR family of regulatory genes

“…The transcription start site of the malR promoter coincides with the first G of the predicted GTG translational start codon, and thus the malR transcript lacks a conventional untranslated leader sequence and ribosome-binding site, consistent with the absence of a purine-rich Shine-Dalgarno sequence complementary to the 3Ј end of the 16S RNA upstream of the malR-coding region. Although several streptomycete mRNAs lack untranslated leader sequences (Janssen, 1993;Strohl, 1992), the malR transcript appears to be only the second example of a leaderless mRNA involved in primary metabolism, the other being that derived from the histidase gene of Streptomyces griseus (Wu et al, 1995).…”

Regulation of the Histidine Utilization (Hut) System in Bacteria

“…Inactivation occurs upon induction of sporulation, nutritional downshift, or phosphate limitation (77). Urocanate may also play a role in this regulation, but its role is complicated by the fact that there is disagreement about whether urocanate is necessary for transcription of hutH (78,168). The nature of the factor(s) that activates and inactivates the histidase of S. griseus is unknown.…”

“…The identity of the hutH gene has been established (167). The hutH gene is unlinked to any other hut gene and is transcribed as a single-gene operon (168). Although early reports argue that hutH transcription is constitutive (78), it now appears that hutH is transcriptionally regulated by a repressor, since the presence of high-copy-number clones of the noncoding region upstream from hutH causes hutH to be derepressed (168).…”

“…The hutH gene is unlinked to any other hut gene and is transcribed as a single-gene operon (168). Although early reports argue that hutH transcription is constitutive (78), it now appears that hutH is transcriptionally regulated by a repressor, since the presence of high-copy-number clones of the noncoding region upstream from hutH causes hutH to be derepressed (168). Current annotation suggests that the remaining hut genes, though unlinked to hutH, may form a single operon elsewhere on the Streptomyces coelicolor chromosome (http://www.ecocyc.org/).…”

Expression of a streptomycete leaderless mRNA encoding chloramphenicol acetyltransferase in Escherichia coli

“…While genes that encode leaderless mRNA are relatively rare, more than 30 have been identified (11,23,32) since the Escherichia coli phage cI repressor was first reported in 1976 (22). Observations of leaderless mRNA (11) in Bacteria, Archaea, Eucarya, and eucaryotic organelles suggest that sequence and/or structural information contained within the coding sequence are sufficient to signal the translational start site and reading frame in these diverse biological systems.…”