Improved hydrogen production with expression of hemH and lba genes in chloroplast of Chlamydomonas reinhardtii

The coding region of both the ferrochelatase gene, hemH, from Bradyrhizobium japonicum, and the leghemoglobin gene, lba, from Glycine max, were transferred into chloroplast of Chlamydomonas reinhardtii. As a result, transgenic C. reinhardtii cultures more rapidly consumed O2 and increased H2 output compared with controls in both sulfur-free and sulfur-containing medium. H2 production of the transgenic algal cultures in sulfur-free medium was 4-fold greater than that of control cultures, ∼3.3 ml bottle−1. Maximum expression of the hemH-lba fusion protein on day 5 coincided with the lowest levels of O2 content and the highest H2 evolution rate detected over 7 days of anaerobic induction in sulfur-free medium. When the concentration of sulfate in the growth medium was restored to 12.5 or 50 μM, O2 consumption and H2 yield decreased more slowly in the transgenic algal cultures than in the control cultures. These results demonstrate that expression of the hemH-lba fusion protein in chloroplast of C. reinhardtii improved their H2 yield by decreasing O2 content in the medium, thereby representing the potential for H2 production in green algae to be improved.