Poster: Posttranscriptional Regulation

Abs # 1080: Analysis of the proteloytic processing of the Chlamydomonas reinhardii protein, periplasmic carbonic anhydrase (pCA).

Presenter:	Wagner, Ryan L, rywagner@iastate.edu
Authors	Wagner, Ryan L (A)   Spalding, Martin H (A)
Affiliations:	(A): Iowa State University

The Chlamydomonas reinhardii gene, CAH1 encodes a secreted periplasmic carbonic anhydrase protein that undergoes distinct proteolytic processing post-translationally. The active periplasmic protein, pCA1, is a holoprotein comprised of two different subunits. Interestingly, the open reading frame of the CAH1 gene contains the complete sequences for a leader sequence, the two subunits (a large subunit and small subunit) and a connecting spacer region between the large and small subunits. The processed protein lacks the leader and the spacer sequences indicating that these distinct regions are removed via post-translational processing. In addition to the proteolytic processing, the protein is a known glycoprotein, indicating additional post-translational processing occurs. The occurrence of post-translational proteolytic processing of a secretory protein in Chlamydomonas may indicate the involvement of endomembrane system proteases such as the Kex2-related proteases. In order to elucidate the post-translational mechanism utilized to process the pCA1 protein, we have developed a series of CAH1 mutants for expression and analysis in the model plant system, Arabidopsis. Site directed mutagenesis was used to disrupt potentially conserved or critical sites at the 5' and 3' termini of the CAH1 spacer sequence. We report here the development of the site directed mutant constructs, and the subsequent effect on pCA1 processing. Identification of the conserved regions of proteolytic processing may provide insight into the identity and/or action of protease(s) involved in the post-translational processing of pCA1.