Effects of the Photosystem II Inhibitors CCCP and DCMU on Hydrogen Production by the Unicellular Halotolerant Cyanobacterium<i>Aphanothece halophytica</i>

Abstract

The unicellular halotolerant cyanobacterium Aphanothece halophytica is a potential dark fermentative producer of molecular hydrogen (H 2 ) that produces very little H 2 under illumination. One factor limiting the H 2 photoproduction of this cyanobacterium is an inhibition of bidirectional hydrogenase activity by oxygen (O 2 ) obtained from splitting water molecules via photosystem II activity. The present study aimed to investigate the effects of the photosystem II inhibitors carbonyl cyanide m-chlorophenyl hydrazone (CCCP) and 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU) on H 2 production of A . halophytica under light and dark conditions and on photosynthetic and respiratory activities. The results showed that A . halophytica treated with CCCP and DCMU produced H 2 at three to five times the rate of untreated cells, when exposed to light. The highest H 2 photoproduction rates,<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" id="M1"><mml:mn fontstyle="italic">2</mml:mn><mml:mo>.</mml:mo><mml:mn fontstyle="italic">26</mml:mn><mml:mo> </mml:mo><mml:mo>±</mml:mo><mml:mo> </mml:mo><mml:mn fontstyle="italic">0</mml:mn><mml:mo>.</mml:mo><mml:mn fontstyle="italic">24</mml:mn></mml:math>and<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" id="M2"><mml:mn fontstyle="italic">3</mml:mn><mml:mo>.</mml:mo><mml:mn fontstyle="italic">63</mml:mn><mml:mo> </mml:mo><mml:mo>±</mml:mo><mml:mo> </mml:mo><mml:mn fontstyle="italic">0</mml:mn><mml:mo>.</mml:mo><mml:mn fontstyle="italic">26</mml:mn><mml:mo> </mml:mo><mml:mo> </mml:mo></mml:math> μ mol H 2 g −1 dry weight h −1 , were found in cells treated with 0.5 μ M CCCP and 50 μ M DCMU, respectively. Without inhibitor treatment, A . halophytica incubated in the dark showed a significant increase in H 2 production compared with cells that were incubated in the light. Only CCCP treatment increased H 2 production of A . halophytica during dark incubation, because CCCP functions as an uncoupling agent of oxidative phosphorylation. The highest dark fermentative H 2 production rate of<mml:math xmlns:mml="http://www.w3.org/1998/Math/MathML" id="M3"><mml:mn fontstyle="italic">39</mml:mn><mml:mo>.</mml:mo><mml:mn fontstyle="italic">50</mml:mn><mml:mo> </mml:mo><mml:mo>±</mml:mo><mml:mo> </mml:mo><mml:mn fontstyle="italic">2</mml:mn><mml:mo>.</mml:mo><mml:mn fontstyle="italic">13</mml:mn><mml:mo> </mml:mo><mml:mo> </mml:mo></mml:math> μ mol H 2 g −1 dry weight h −1 was found in cells treated with 0.5 μ M CCCP after 2 h of dark incubation. Under illumination, CCCP and DCMU inhibited chlorophyll fluorescence, resulting in a low level of O 2 , which promoted bidirectional hydrogenase activity in A . halophytica cells. In addition, only CCCP enhanced the respiration rate, further reducing the O 2 level. In contrast, DCMU reduced the respiration rate in A . halophytica.