There are four forms of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) found in nature. Forms I, II, and III catalyse the carboxylation and oxygenation of ribulose 1,5-bisphosphate, while form IV, also called the Rubisco-like protein (RLP), does not catalyse either of these reactions. There appear to be six different clades of RLP. Although related to bona fide Rubisco proteins at the primary sequence and tertiary structure levels, RLP from two of these clades is known to perform other functions in the cell. Forms I, II, and III Rubisco, along with form IV (RLP), are thought to have evolved from a primordial archaeal Rubisco. Structure/function studies with both archaeal form III (methanogen) and form I (cyanobacterial) Rubisco have identified residues that appear to be specifically involved with interactions with molecular oxygen. A specific region of all form I, II, and III Rubisco was identified as being important for these interactions.

Ribulose 1, 5-bisphosphate (RuBP) carboxylase/oxygenase (Rubisco) is a globally significant biocatalyst that facilitates the removal and sequestration of CO 2 from the biosphere. Rubiscocatalyzed CO 2 reduction thus provides virtually all the organic carbon utilized by living organisms. Despite catalyzing the rate-limiting step of photosynthetic and chemoautotrophic CO 2 assimilation, Rubisco is markedly inefficient as the competition between O 2 and CO 2 for the same substrate limits the ability of aerobic organisms to obtain maximum amounts of organic carbon for CO 2 -dependent growth. Random and site-directed mutagenesis procedures were coupled with genetic selection to identify an "oxygen insensitive" mutant cyanobacterial (Synechococcus sp. strain PCC 6301) Rubisco that allowed for CO 2 -dependent growth of a host bacterium at an oxygen concentration that inhibited growth of the host containing wild-type Synechococcus Rubisco. The mutant substitution, A375V, was identified as an intragenic suppressor of D103V, a negative mutant enzyme incapable of supporting autotrophic growth. Ala-375 (Ala-378 of spinach Rubisco) is a conserved residue in all form I (plant-like) Rubiscos. Structure-function analyses indicate that the A375V substitution decreased the enzyme's oxygen sensitivity (and not CO 2 /O 2 specificity), possibly by rearranging a network of interactions in a fairly conserved hydrophobic pocket near the active site. These studies point to the potential of engineering plants and other significant aerobic organisms to fix CO 2 unfettered by the presence of O 2 .The Calvin-Benson-Bassham (CBB) reductive pentose phosphate pathway provides a way for many organisms to reduce carbon dioxide to organic carbon, a process that is vital for life on earth (1). Rubisco is the rate-limiting enzyme in this pathway, catalyzing the initial steps in both autotrophic carbon assimilation (CO 2 fixation or carboxylation) and photooxidative metabolism (O 2 fixation or oxygenation) via parallel reaction mechanisms that share a common acceptor intermediate, the enediol form of RuBP (2). Thus, in aerobic organisms, the promiscuity of this enediolate for both CO 2 and O 2 limits autotrophic CO 2 assimilation. Severe limitations imposed on the enzyme's efficiency, primarily due to the competition between CO 2 and O 2 at the same active site and the poor turnover rate, have prompted numerous studies directed towards improving the enzyme's net carboxylation efficiency (2). Thus, an increase

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