Abstract: 

The marine cyanobacterium Prochlorococcus fixes CO2 in the Calvin-Benson-Bassham (CBB) cycle during the day and shifts to the pentose phosphate pathway (PPP) for nucleotide synthesis at night. The intrinsically disordered protein CP12 is known to form a regulatory complex with two key CBB enzymes, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and phosphoribulokinase (PRK), to inhibit their activities. However, it is unreported whether this mechanism exists and what the structure of its protein complex looks like in Prochlorococcus. Cyanophages that infect Prochlorococcus also encode and express the CP12 protein, lacking other CBB-related genes. Our previous work found that the cyanophage-encoded CP12 can replace host CP12 and bind to host GAPDH, thereby participating in the host Calvin cycle during the infection process.
In this study, I employed protein crosslinking, rapid protein liquid chromatography, Grafix, transmission electron microscopy, and cryo-electron microscopy to investigate the specific structure of the protein complex mediated by CP12 in MED4 under dark conditions and during P-HM2 infection. It was found that the native protein complex was unstable in the dark condition. Direct glutaraldehyde crosslinking stabilized the complex and yielded particles resembling previously reported structures but increased conformational heterogeneity. GraFix crosslinking improved particle uniformity and produced a reconstructed structure consistent with the expected size. Under the P-HM2 infection condition, phage-encoded CP12 could replace the host CP12. However, the infected complex showed increased aggregation and larger particle size, indicating that the phage CP12 might alter the surface properties of the complex. 
This study provides the first structural-level evidence that despite lacking the N-terminal cysteine residues and the AWD_VEEL domains, MED4 CP12 can form an unstable trimeric complex in vivo, and the phage CP12 can functionally replace the host CP12 during infection.