Molecular and biochemical analysis of the α class carbonic anhydrases in Caenorhabditis elegans

Mol Biol Rep. 2011 Mar;38(3):1777-85. doi: 10.1007/s11033-010-0292-y. Epub 2010 Sep 15.

Abstract

In this study, in silico analysis of the Caenorhabditis elegans genome revealed six genes (cah-1, cah-2, cah-3, cah-4, cah-5, and cah-6) possibly encoding α class CAs (carbonic anhydrase). Real-time RT-PCR analysis revealed the temporal expression pattern of each gene, as well as changes in expression levels under different atmospheric conditions (stress). Cah-3 and cah-4 showed the highest levels of transcript accumulation, while most genes responded to the stress conditions. Yeast complementation showed that cah-3 was able to complement the function of Saccharomyces cerevisiae CA (NCE103) in vivo. Recombinant CAH-3, CAH-4a and CAH-5 enzymes, expressed in Escherichia coli were used for in vitro measurement of CA activity. However, in vitro activity was only detectable for CAH-4a. RNAi by feeding was performed on wild-type C. elegans for all genes. The worms were examined for a visible phenotype under normal and stress conditions (pH, CO(2)/O(2)). Silencing cah-3 and cah-4 may reduce the life-span of the worms (at 22 °C).

MeSH terms

  • Animals
  • Caenorhabditis elegans / enzymology*
  • Caenorhabditis elegans / genetics
  • Caenorhabditis elegans / growth & development
  • Caenorhabditis elegans Proteins / chemistry
  • Caenorhabditis elegans Proteins / genetics*
  • Caenorhabditis elegans Proteins / metabolism*
  • Carbonic Anhydrases / chemistry
  • Carbonic Anhydrases / genetics*
  • Carbonic Anhydrases / metabolism*
  • Electrophoresis, Polyacrylamide Gel
  • Enzyme Assays
  • Gene Expression Regulation, Developmental
  • Gene Expression Regulation, Enzymologic
  • Genetic Complementation Test
  • Longevity
  • RNA Interference
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Saccharomyces cerevisiae / genetics
  • Sequence Alignment
  • Sequence Analysis, Protein

Substances

  • Caenorhabditis elegans Proteins
  • RNA, Messenger
  • Carbonic Anhydrases