Structure and identification of a pterin dehydratase-like protein as a RuBisCO assembly factor in the alpha-carboxysome [Plant Biology]

January 23rd, 2014 by Wheatley, N. M., Sundberg, C. D., Gidaniyan, S. D., Cascio, D., Yeates, T. O.

Carboxysomes are proteinaceous bacterial microcompartments that increase the efficiency of the rate-limiting step in carbon fixation by sequestering reaction substrates. Typically, α-carboxysomes are genetically encoded as a single operon expressing the microcompartment's structural proteins and its encapsulated enzymes. In addition, depending on phylogeny, as many as thirteen other genes are found to co-occur near or within α-carboxysome operons. One of these genes codes for a protein with distant homology to pterin-4a-carbinolamine dehydratase (PCD) enzymes. It is present in all α-carboxysome containing bacteria and has homologs in algae and higher plants. Canonical PCDs play an important role in amino acid hydroxylation, a reaction not associated with carbon fixation. We determined the crystal structure of an α-carboxysome PCD-like protein from the chemoautotrophic bacterium Thiomonas intermedia K12, at a resolution of 1.3 Å. The protein retains a three-dimensional fold similar to canonical PCDs, though the prominent active site cleft present in PCD enzymes is disrupted in the α-carboxysome PCD-like protein. Using a cell-based complementation assay, we tested the PCD-like proteins from T. intermedia and two additional bacteria, and found no evidence for PCD enzymatic activity. However, we discovered that heterologous co-expression of the PCD-like protein from H. neapolitanus with RuBisCO and GroELS in E. coli increased the amount of soluble, assembled RuBisCO recovered from cell lysates compared to co-expression of RuBisCO with GroELS alone. We conclude that this conserved PCD-like protein, renamed here alpha carboxysome RuBisCO assembly factor (or acRAF), is a novel RuBisCO chaperone integral to α-carboxysome function.