The Regulation of Bacteriochlorophyll Biosynthesis by the PufQ Protein of Rhodobacter Capsulatus

Abstract

The first gene of the puf operon of Rhodobacter capsulatus (pufQ), is thought to be involved in the regulation of bacteriochlorophyll (Bchl) biosynthesis [I]. A puf operon deletion mutant of R. capsulatus (ΔRC6) could only form very small amounts of Bchl (present in vivo as LHII), although transcription of the puc operon was not affected 121. When the pufQ gene was added to ΔRC6 in trans, LHII was formed at near normal levels, but the transconjugants could not grow photosynthetically since they lacked genes for the RC and LHI [1,2]. It has also been shown that seven mutants of R. capsulatus each deficient in a different bch gene were each unable to synthesize an intermediate of Bchl biosynthesis under low oxygen conditions ranging all the way from protoporphyrin IX (Proto) to bacteriochlorophyllide after the puf operon had been deleted [1]. Addition of the pufQ gene in trans restored the synthesis of the intermediate in each case. Nevertheless, it was found that mRNA from at least two bch genes was induced normally by low oxygen in a mutant of R. capsulatus which was unable to induce transcription of the pufQ gene [3]. While PufQ may not be involved in regulation of bch genes encoding enzymes for the latter part of the tetrapyrrole pathway, it may still be involved in regulation of one or more hem genes encoding enzymes catalyzing the biosynthesis of Proto during the early part of the pathway [4]. Lowering the oxygen tension of the medium in R. capsulatus resulted in a 2- to 3-fold increase in promoter activity for hemA, encoding δ-aminolevulinic acid (ALA) synthase, which was similar to the increase observed in transcription rates for several bch genes [5]. As well, activity for the second step of Proto synthesis, porphobilinogen (PBG) synthase encoded by hemB, was reported to be regulated by the oxygen tension of the medium [6]. We also found activity for one or both of the enzymes constituting PBG deaminase activity, hydroxymethylbilane (HMB) synthase encoded by hemC and/or uroporphyrinogen Ill (Urogen) synthase encoded by hemD, to be induced by low oxygen [4]. This induction did not occur in the puf operon deletion mutant, ΔRC6, but was restored by the addition of the pufQ gene in trans in strain ΔRC6(pΔ4) [4]. Transcription of hemE which encodes Urogen decarboxylase is apparently not regulated by oxygen tensions, however [7]. We have prepared a mutant (JSΔQ) which lacks only the pufQ gene and have found that restoration of the pufQ gene in trans restored not only Bchl synthesis, but also photosynthetic competence. We report herein the analysis of ALA synthase, PBG synthase, and PBG deaminase activities under both aerobic and semiaerobic conditions in the JSΔQ mutant of R. capsulatus in comparison with those found in the wild-type strain grown under similar conditions.