Proteomic analysis of mutant cell cultures for two mitchondrial carbonic anhydrases like genes

VILLARREAL, F;; MARTIN, V,; SCHROTT, G, ; ZABALETA, E

Carlos Paz, Cordoba, Argentina

Congreso; XLIV, reunion anual - Sociedad argentina de investigación en Bioquímica y Biología Molecular (SAIB); 2008

<!-- /* Style Definitions */ p.MsoNormal, li.MsoNormal, div.MsoNormal {mso-style-parent:""; margin:0cm; margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:12.0pt; font-family:"Times New Roman"; mso-fareast-font-family:"Times New Roman";} @page Section1 {size:595.3pt 841.9pt; margin:70.85pt 3.0cm 70.85pt 3.0cm; mso-header-margin:35.4pt; mso-footer-margin:35.4pt; mso-paper-source:0;} div.Section1 {page:Section1;} --> The gene family of putative gamma-class Carbonic Anhydrases in Arabidopsis genome incluyes five members. Three of these genes (CA1, CA2 and CA3), share higher homology with the highly characterizaed gamma-CAs from the archea M. Thermophila, and two CA-Like genes (CAL1 and CAL2) show a minor degree of homology. By bioinformatic analysis, the CALs proteins lacks a number of aminoacid important to create the funcional active site of the mature homotrimer, but they can form part of a mature heterotrimer together with two CA subunits. The five CA and CAL proteins are located into mitochondria and bounded to mitochondrial complex I, forming an extra domain exclusive of photosyntetic organisms. By proteomics analysis, we showed that ca2 mutant cell culture exhibited a 80% reduction in complex I levels, but this feature was not observed in ca1 or ca3 mutants. In this work, we analised the mitochondrial proteome of cal1 mutants and cal1cal2 mutant cell cultures (despite all efforts, we were not able to obtain cal2 cell culture). We observed that cal1 and cal1cal2 cell cultures present a normal amounts of all mitochondrial complexes. A detalled analysis showed that both mutants lacked the same two bands in silver stained 2D BN-SDS gel and only one band in western blots. This suggested an epistatic relationship between CAL1 y CAL2, but this need further investigation.