Researchers discover how photosynthetic organisms regulate and synthesize ATP

Photo credit: Tokyo Tech

ATP, the compound essential for the functioning of photosynthetic organisms such as plants, algae and cyanobacteria, is made by an enzyme called “chloroplast ATP synthase” (CFOCF1). To control ATP production under varying light conditions, the enzyme uses a redox regulatory mechanism that modifies ATP synthetic activity in response to changes in the redox state of cysteine ​​(Cys) residues, which exist as dithiols under reducing (light) conditions , but form a disulfide bond under oxidizing (dark) conditions. However, this mechanism is not yet fully understood.

Well, in a study published in the Proceedings of the National Academy of Sciencesa team of researchers from Japan led by Prof. Toru Hisabori of the Tokyo Institute of Technology (Tokyo Tech) has revealed the role of the amino acid sequences present in CFOCF1showing how the enzyme regulates ATP production in photosynthetic organisms.

Understand what is the conformation of the amino acids present in CFOCF1 contributes to the redox regulation mechanism, the researchers used the unicellular green alga Chlamydomonas reinhardtii to produce the enzyme. “By using the powerful genetics of Chlamydomonas reinhardtii as a model organism for photosynthesis, we performed a comprehensive biochemical analysis of CFOCF1 molecule,” explains Prof. Hisabori.

Using the alga as the host organism, the team introduced plasmids (extrachromosomal DNA molecules that can replicate independently) specific to the F1 Part of the CFOCF1 Protein, namely the part of the enzyme that contains catalytic sites for ATP synthesis. They also introduced mutant versions of the gene to alter the protein’s amino acid sequences, specifically targeting the DDE motif (a cluster of negatively charged amino acids), the redox loop, and the β-hairpin domain.

Then they cleaned CFOCF1five different variations of which were generated, containing a wild-type strain with no changes to the amino acid sequence and four mutant strains: one with the DDE motif replaced with neutral amino acids, Asn-Asn-Gln, one without the β-hairpin domain, one without the redox loop and one lacking both the redox loop and the β-hairpin domain.

When testing the ATP synthesis activity of these mutants under reducing (mimicking light conditions) and oxidizing (mimicking dark conditions) conditions, the researchers found that the wild-type enzyme and the mutant enzyme with changes in the DDE motif functioned normally (high activity shown in reduction and low activity in oxidation). However, the enzyme complexes lacking the redox loop or the β-hairpin domain showed no redox response, indicating that both regions were involved in the redox regulation mechanism.

The researchers proposed that under dark conditions, the disulfide bond between the Cys residues makes the redox loop rigid and weakens the interaction between the redox loop and the β-hairpin. This causes the β-hairpin to get stuck in a cavity in the protein. However, when the disulfide bond is reduced in the presence of light, the redox loop regains its flexibility and pulls the β-hairpin out of the cavity, allowing it to participate in ATP synthesis activity.

“The redox regulation of ATP synthesis is achieved through a cooperative interaction between two domains of the γ subunit of CFOCF1 unique to photosynthetic organisms,” says Prof. Hisabori. “We propose that this results from the interaction of the β-hairpin and the redox loop with the catalytic site.”

The results are an important step towards a better understanding of the photosynthetic process with the potential for significant implications for agriculture and bioenergy.

More information:
Kentaro Akiyama et al., Two specific domains of the γ-subunit of the chloroplast F o F 1 provide redox regulation of ATP synthesis through conformational changes, Proceedings of the National Academy of Sciences (2023). DOI: 10.1073/pnas.2218187120

Provided by Tokyo Institute of Technology

Citation: Researchers Reveal How Photosynthetic Organisms Regulate and Synthesize ATP (2023 February 22) Retrieved February 22, 2023 from

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