Kinetic and physical properties of a recombinant RuBisCO from a chemoautotrophic endosymbiont.
Ribulose 1,5 bisphosphate carboxylase/oxygenase (RuBisCO), which catalyzes the key step in autotrophic CO2 fixation, is present at high activity in the symbiont-containing tissues of many hydrothermal vent invertebrates. The genes encoding RuBisCO from a gill endosymbiont of the hydrothermal vent gastropod Alviniconcha hessleri have been cloned, sequenced, and functionally expressed in Escherichia coli under control of the lac promoter in the vector pBlueScript. The purified protein is a hexadecamer (L8S8) with an apparent molecular weight of 554 kDa and a specific carboxylase activity of 2.9 mumol/min/mg protein. Unlike previously characterized RuBisCOs, which display sharp temperature optima, the symbiont RuBisCO maintains a high level of activity over a broad temperature span, although it is not thermally stable after extended exposure to temperatures above 50 degrees C. This funding suggests an adaptation to the thermal transients measured at vent openings where the host snail resides. The enzyme also maintains 90% of its 1 atm activity at 370 atm pressure, whereas spinach RuBisCO retains 28% activity under similar conditions. At 1 atm pressure and 20 degrees C, the Km(CO2) and the relative substrate specificity (Srel) of the symbiont enzyme were 80 and 32.5 mumol/L, respectively, which are similar to values reported for RuBisCOs from cyanobacteria and the purple photosynthetic bacteria. The relatively low specificity of the enzyme for substrate CO2 indicates that the intracellular environment of the endosymbionts may be microaerophilic for RuBisCO to maintain net carboxylation.
Stein
J L
JL
Marine Biology Research Division, Scripps Institution of Oceanography, La Jolla, CA 92093.
Felbeck
H
H
D000818:Animals; D001419:Bacteria / Q000201:enzymology*; D004795:Enzyme Stability; D011994:Recombinant Proteins / Q000378:metabolism; D012273:Ribulose-Bisphosphate Carboxylase / Q000235:genetics / Q000378:metabolism*; D012908:Snails / Q000382:microbiology*; D013559:Symbiosis
D011994:Recombinant Proteins; D012273:Ribulose-Bisphosphate Carboxylase