
%0 Journal Article
%A So, Anthony K.-C. 
%A Cot, Swan S.-W. 
%A Espie, George S. 
%T Characterization of the C-terminal extension of carboxysomal carbonic anhydrase from <emph type="2">Synechocystis</emph> sp. PCC6803
%J Functional Plant Biology
%D 2002/03/19
%V 29
%N 3
%P 183-194
%K carbonic anhydrase, carboxysome, CO2, COS, C-terminal extension, cyanobacteria, mass spectrometry, protein interaction, Synechocystis.
%R https://doi.org/10.1071/PP01179
%U https://www.publish.csiro.au/paper/PP01179 
%X Sequence analysis of the carboxysomal carbonic anhydrase (CcaA) from Synechocystis PCC6803, Synechococcus PCC7942 and Nostoc ATCC29133, indicated high sequence identity to the &#946; class of plant and bacterial carbonic anhydrases (CA), and conservation of the active site region. However, the cyanobacterial enzyme has a C-terminal extension of about 75 amino acids (aa) not found in the plant enzymes, and largely absent from other bacterial enzymes. Using recombinant DNA technology, genes encoding C-terminal truncation products of up to 127 aa were overexpressed in E. coli, and partially purified lysates were analysed for CA-mediated exchange of 18O between 13C18O2and H216O. Recombinant CcaA proteins with up to 60 aa removed (CcaA&#916;60) were catalytically competent, but beyond this there was an abrupt loss of activity. CcaA&#916;0, along with CcaA&#916;40 and CcaA&#916;60, also catalysed the hydrolysis of carbon oxysulfide (COS; an isoelectronic structural analogue of CO2), but CcaA&#916;63 and CcaA&#916;127 did not, indicating that truncations greater than 62 aa resulted in a general loss of catalytic competency. Analysis of protein-protein interaction using the yeast two-hybrid system revealed that CcaA did not interact with the large or small Rubisco subunits (RbcL and RbcS, respectively) of Synechocystis, but there was strong CcaA-CcaA interaction. This protein interaction also ceased with C-terminal truncations in CcaA greater than 60 aa. The correlation between loss of CcaA-CcaA interaction and CcaA catalytic activity suggests that the proximal portion of the C-terminal extension is required for oligomerization, and that this oligomerization is essential for catalysis by the cyanobacterial enzyme. Thus, the C-terminal extension may play an important role in the function of CA within cyanobacterial carboxysomes, which is not required by the higher plant enzymes. 
