A plant-type (beta-class) carbonic anhydrase in the thermophilic methanoarchaeon Methanobacterium thermoautotrophicum.
Carbonic anhydrase, a zinc enzyme catalyzing the interconversion of carbon dioxide and bicarbonate, is nearly ubiquitous in the tissues of highly evolved eukaryotes. Here we report on the first known plant-type (beta-class) carbonic anhydrase in the archaea. The Methanobacterium thermoautotrophicum DeltaH cab gene was hyperexpressed in Escherichia coli, and the heterologously produced protein was purified 13-fold to apparent homogeneity. The enzyme, designated Cab, is thermostable at temperatures up to 75 degrees C. No esterase activity was detected with p-phenylacetate as the substrate. The enzyme is an apparent tetramer containing approximately one zinc per subunit, as determined by plasma emission spectroscopy. Cab has a CO(2) hydration activity with a k(cat) of 1.7 x 10(4) s(-1) and K(m) for CO(2) of 2.9 mM at pH 8.5 and 25 degrees C. Western blot analysis indicates that Cab (beta class) is expressed in M. thermoautotrophicum; moreover, a protein cross-reacting to antiserum raised against the gamma carbonic anhydrase from Methanosarcina thermophila was detected. These results show that beta-class carbonic anhydrases extend not only into the Archaea domain but also into the thermophilic prokaryotes.
Smith
K S
KS
Department of Biochemistry, The Pennsylvania State University, University Park, Pennsylvania 16802, USA.
Ferry
J G
JG
D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D002245:Carbon Dioxide / Q000378:metabolism; D002257:Carbonic Anhydrase Inhibitors; D002256:Carbonic Anhydrases / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D006358:Hot Temperature; D017014:Methanobacterium / Q000201:enzymology* / Q000235:genetics; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000235:genetics; D011994:Recombinant Proteins / Q000145:classification / Q000302:isolation & purification; D017386:Sequence Homology, Amino Acid; D014867:Water / Q000378:metabolism
D019843:Archaeal Proteins; D002257:Carbonic Anhydrase Inhibitors; D010940:Plant Proteins; D011994:Recombinant Proteins; D014867:Water; D002245:Carbon Dioxide; D002256:Carbonic Anhydrases