Differential regulation of the Fe-hydrogenase during anaerobic adaptation in the green alga Chlamydomonas reinhardtii.
Chlamydomonas reinhardtii, a unicellular green alga, contains a hydrogenase enzyme, which is induced by anaerobic adaptation of the cells. Using the suppression subtractive hybridization (SSH) approach, the differential expression of genes under anaerobiosis was analyzed. A PCR fragment with similarity to the genes of bacterial Fe-hydrogenases was isolated and used to screen an anaerobic cDNA expression library of C. reinhardtii. The cDNA sequence of hydA contains a 1494-bp ORF encoding a protein with an apparent molecular mass of 53.1 kDa. The transcription of the hydrogenase gene is very rapidly induced during anaerobic adaptation of the cells. The deduced amino-acid sequence corresponds to two polypeptide sequences determined by sequence analysis of the isolated native protein. The Fe-hydrogenase contains a short transit peptide of 56 amino acids, which routes the hydrogenase to the chloroplast stroma. The isolated protein belongs to a new class of Fe-hydrogenases. All four cysteine residues and 12 other amino acids, which are strictly conserved in the active site (H-cluster) of Fe-hydrogenases, have been identified. The N-terminus of the C. reinhardtii protein is markedly truncated compared to other non-algal Fe-hydrogenases. Further conserved cysteines that coordinate additional Fe-S-cluster in other Fe-hydrogenases are missing. Ferredoxin PetF, the natural electron donor, links the hydrogenase from C. reinhardtii to the photosynthetic electron transport chain. The hydrogenase enables the survival of the green algae under anaerobic conditions by transferring the electrons from reducing equivalents to the enzyme.
Happe
Thomas
T
Botanisches Institut der Universität Bonn, Germany. t.happe@uni-bonn.de
Kaminski
Annette
A
D000222:Adaptation, Physiological / Q000235:genetics*; D000595:Amino Acid Sequence; D000693:Anaerobiosis; D000818:Animals; D001483:Base Sequence; D015152:Blotting, Northern; D016825:Chlamydomonas reinhardtii / Q000378:metabolism* / Q000502:physiology; D003001:Cloning, Molecular; D018076:DNA, Complementary; D004926:Escherichia coli / Q000235:genetics; D015971:Gene Expression Regulation, Enzymologic; D006864:Hydrogenase / Q000235:genetics* / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
D018076:DNA, Complementary; D007506:Iron-Sulfur Proteins; D011994:Recombinant Proteins; C118830:iron hydrogenase; D006864:Hydrogenase