SummaryPyruvate formate-lyase (PFL) catalyses the non-oxidative dissimilation of pyruvate to formate and acetylCoA using a radical-chemical mechanism. The enzyme is enzymically interconverted between inactive and active forms, the active form contains an organic free radical located on a glycyl residue in the C-terminal portion of the polypeptide chain. Introduction of the radical into PFL only occurs anaerobically, and the activating enzyme responsible is an iron-sulphur protein that uses S-adenosyl methionine as cofactor and reduced flavodoxin as reductant. As the radical form of PFL is inactivated by molecular oxygen it is safeguarded during the transition to aerobiosis by conversion back to the radical-free, oxygen-stable form. This reaction is catalysed by the anaerobically induced multimeric enzyme alcohol dehydrogenase. The genes encoding PFL and its activating enzyme are adjacent on the chromosome but form discrete transcriptional units. This genetic organization is highly conserved in many, but not all, organisms that have PFL. Recent studies have shown that proteins exhibiting significant similarity to PFL and its activating enzyme are relatively widespread in facultative and obligate anaerobic eubacteria, as well as archaea. The physiological function of many of these PFL-like enzymes remains to be established. It is becoming increasingly apparent that glycyl radical enzymes are more prevalent than previously surmised. They represent a class of enzymes with unusual biochemistry and probably predate the appearance of molecular oxygen.

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Ribulose 1,5-bisphosphate carboxylase/oxygenase (RubisCO) catalyzes the key reaction of the Calvin reductive pentose phosphate cycle and as such is responsible for life as we know it. This enzyme has been intensively studied for decades. Evidence that RubisCO phylogenies are incongruent with those derived from other macromolecules has been accumulating and recent discoveries have driven home this point. Here we review findings regarding RubisCO phylogeny and discuss these in the context of the important biochemical and structural features of the enzyme. The implications for the engineering of improved RubisCO enzymes are considered.

The factors affecting the regulation of photosynthetic carbon fixation in diverse phytoplankton populations are not yet understood. To this end, we have measured the expression of the gene (rbcL) for the major carbon fixation enzyme, ribulose-1,5-bisphosphate carboxylase/oxygenase, in coastal phytoplankton populations off Cape Hatteras and in oligotrophic oceanic picoplankton near Bermuda. Using gene probes specific for the cyanobacterial/chlorophytic clade and the chromophytic clade (diatoms, chrysophytes, prymnesiophytes, and others) of Form I rbcL genes (''cyano'' and ''chromo'' probes, respectively), we have measured rbcL messenger ribonucleic acid (mRNA) levels in size-fractionated coastal waters, in a decktop diel incubator and a Lagrangian drifter study, and in vertical profiles in stratified, oligotrophic ocean water. In coastal waters influenced by estuarine plumes, an equal distribution of carbon fixation between the picoplankton and the micro/nannoplankton occurred, with cyano rbcL mRNA coinciding with Synechococcus counts in the Ͻ1-m fraction, with the majority of the chromo rbcL mRNA expression occurring in the larger sized phytoplankton fraction. In profiles of oligotrophic oceanic waters, the cyano rbcL mRNA was found in the upper water column (ϳ50-m depth) and coincided with peaks in Synechococcus counts. The chromo rbcL mRNA was concentrated at the subsurface chlorophyll a (Chl a) maximum (ϳ85 m) and corresponded to red-fluorescing cell counts, thought to be picoeucaryotes and diatoms. Photosynthetic carbon fixation and RUBISCO enzyme activity encompassed both cyano and chromo rbcL mRNA peaks, suggesting a near equal contribution to carbon fixation in the water column by these two phytoplankton clades. Both decktop diel incubator studies and a Lagrangian drifter study in coastal waters indicated cyano rbcL transcription in the morning and chromophytic rbcL transcription in the late afternoon/early evening. Thus, the two major clades of RUBISCOcontaining phytoplankton occupy separate niches in time, space, and cell size in the waters off Cape Hatteras. The factors determining such clade-specific niches may include efficiency of nutrient utilization, differences in relative carboxylase/oxygenase activity ( values) of cyano (Form IB) and chromophytic (Form ID) RUBISCOs, and differences in pigment composition/adaptation to light regimes. Additionally, we propose that chromo rbcL mRNA may be indicative of new production, whereas cyano rbcL mRNA correlates with recycled production in stratified, oligotrophic oceanic environments.Although photosynthetic carbon fixation is one of the most important biological processes occurring in the ocean, little is known concerning the molecular regulation of this process in natural phytoplankton assemblages. The enzyme responsible for the vast majority of photosynthetic carbon fixation by phytoplankton is ribulose bisphosphate carboxylase/oxygenase (RUBISCO) (Raven 1993). There is evidence for ␤-carboxylation (Descolas- Gros and Fontugne 1985;Colman 1989;C...