Chlorophyll a is a favorable substrate for Chlamydomonas Mg-dechelatase encoded by STAY-GREEN

Abstract

Mg removal from chlorophyll by Mg-dechelatase is the first step of chlorophyll degradation. Recent studies showed that in Arabidopsis, Stay Green (SGR) encodes Mg-dechelatase. Though the Escherichia coli expression system is advantageous for investigating the properties of Mg-dechelatase, Arabidopsis Mg-dechelatase is not successfully expressed in E. coli. Chlamydomonas reinhardtii SGR (CrSGR) has a long, hydrophilic tail, suggesting that active CrSGR can be expressed in E. coli. After the incubation of chlorophyll a with CrSGR expressed in E. coli, pheophytin a accumulated, indicating that active CrSGR was expressed in E. coli. Substrate specificity of CrSGR against chlorophyll b and an intermediate molecule of the chlorophyll b degradation pathway was examined. CrSGR exhibited no activity against chlorophyll b and low activity against 7-hydroxymethyl chlorophyll a, consistent with the fact that chlorophyll b is degraded only after conversion to chlorophyll a. CrSGR exhibited low activity against divinyl chlorophyll a and chlorophyll a', and no activity against chlorophyllide a, protochlorophyll a, chlorophyll c, and Zn-chlorophyll a. These observations indicate that chlorophyll a is the most favorable substrate for CrSGR. When CrSGR was expressed in Arabidopsis cells, the chlorophyll content decreased, further confirming that SGR has Mg-dechelating activity in chloroplasts.

Mg‐dechelatase is involved in the formation of photosystem II but not in chlorophyll degradation in Chlamydomonas reinhardtii

“…A previous study showed that CrSGR has the same catalytic properties as Arabidopsis SGR (AtSGR)1 and that transiently expressed CrSGR ‐induced Chl degradation in Arabidopsis (Matsuda et al ., ). In this study, CrSGR was constitutively expressed in an Arabidopsis sgr triple mutant using the 35S promoter (CrSGR‐35S) or an Arabidopsis native promoter (2 kbp upstream from the translation start site of AtSGR1 ) (CrSGR‐N) (Figure S7).…”

“…By contrast, Chl b is not present in the core antenna complexes of streptophytes, including Arabidopsis, and SGR can degrade all the Chl in the core antennae, which might be one of the reasons why SGR has different physiological functions between Arabidopsis and Chlamydomonas . It should be noted that AtSGR1 and CrSGR catalyze the same reaction that converts Chl a to Pheo a but not chlorophyllide a to pheophorbide a (Matsuda et al ., ). In addition, CrSGR partly complemented an Arabidopsis sgr triple mutant (Figure S7), and transiently expressed CrSGR in Arabidopsis induced Chl degradation (Matsuda et al ., ).…”

“…It should be noted that AtSGR1 and CrSGR catalyze the same reaction that converts Chl a to Pheo a but not chlorophyllide a to pheophorbide a (Matsuda et al ., ). In addition, CrSGR partly complemented an Arabidopsis sgr triple mutant (Figure S7), and transiently expressed CrSGR in Arabidopsis induced Chl degradation (Matsuda et al ., ). These results indicate that the enzymatic properties between AtSGR and CrSGR are identical, but the physiological functions are different.…”

“…Previously, we showed that Chlamydomonas SGR catalyzes the conversion of Chl a to Pheo a (Matsuda et al, 2016). One possible reason for the reduced F v /F m ratio is a limited supply of Pheo a for PSII formation.…”

“…The Mg in Chl is spontaneously released under acidic conditions (Saga et al ., ), which led to the hypothesis that Pheo a is non‐enzymatically produced from Chl a under such conditions (Christ and Hortensteiner, ). Recently, we identified a Mg‐dechelatase, STAY‐GREEN ( SGR ), in Arabidopsis (Shimoda et al ., ) and Chlamydomonas (Matsuda et al ., ) that removes Mg and converts Chl a to Pheo a . These studies clearly show that Pheo a is not spontaneously produced but is enzymatically synthesized.…”