Phytobilin biosynthesis: cloning and expression of a gene encoding soluble ferredoxin-dependent heme oxygenase from Synechocystis sp. PCC 6803.
The phytobilin chromophores of phycobiliproteins and phytochromes are biosynthesized from heme in a pathway that begins with the opening of the tetrapyrrole macrocycle of protoheme to form biliverdin IX alpha, in a reaction catalyzed by heme oxygenase. A gene containing an open reading frame with a predicted polypeptide that has a sequence similar to that of a conserved region of animal microsomal heme oxygenases was identified in the published genomic sequence of Synechocystis sp. PCC 6803. This gene, named ho1, was cloned and expressed in Escherichia coli under the control of the lacZ promoter. Cells expressing the gene became green colored due to the accumulation of biliverdin IX alpha. The size of the expressed protein was equal to the predicted size of the Synechocystis gene product, named HO1. Heme oxygenase activity was assayed in incubations containing extract of transformed E. coli cells. Incubations containing extract of induced cells, but not those containing extract of uninduced cells, had ferredoxin-dependent heme oxygenase activity. With mesoheme as the substrate, the reaction product was identified as mesobiliverdin IX alpha by spectrophotometry and reverse-phase HPLC. Heme oxygenase activity was not sedimented by centrifugation at 100, 000 g. Expression of HO1 increased several-fold during incubation of the cells for 72 h in iron-deficient medium.
Cornejo
J
J
Division of Biology and Medicine, Brown University, Providence, Rhode Island 02912, USA.
Willows
R D
RD
Beale
S I
SI
D001426:Bacterial Proteins / Q000096:biosynthesis*; D001664:Biliverdine / Q000096:biosynthesis; D003001:Cloning, Molecular; D000458:Cyanobacteria / Q000201:enzymology / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D005288:Ferredoxins; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology*; D005798:Genes, Bacterial / Q000235:genetics; D006419:Heme Oxygenase (Decyclizing) / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D006427:Hemin; D045342:Light-Harvesting Protein Complexes; D008970:Molecular Weight; D010940:Plant Proteins / Q000096:biosynthesis*; D012329:RNA, Bacterial / Q000032:analysis; D012333:RNA, Messenger / Q000032:analysis
D001426:Bacterial Proteins; D005288:Ferredoxins; D045342:Light-Harvesting Protein Complexes; D010940:Plant Proteins; D012329:RNA, Bacterial; D012333:RNA, Messenger; D006427:Hemin; D006419:Heme Oxygenase (Decyclizing); C105245:heme oxygenase-2; D001664:Biliverdine