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In order to elucidate the discrete steps in phosphoenolpyruvate carboxylase (PEPC) evolution concerning Km-PEP and malate tolerance a comparison was made between C3, C3-C4 and C4 species of the dicot genus Flaveria. The PEPCs of this genus are encoded by a gene family comprising three classes: ppcA, ppcB and ppcC [J. Hermans and P. Westhoff(1990) Mol Gen Genet 224:459-468, (1992) Mol Gen Genet 234:275-284]. The ppM of F. trinervia (C4) codes for the C4 PEPC isoform but other plants of the genus contain ppM orthologues too. The C3 plant F. pringlei showed the lowest levels of ppM PEPC mRNA followed by F. pubescens (C3-C4) while the C4-like plant F. brownii displayed RNA amounts close to the C4 species F. trinervia. In contrast to the similar expression profiles of F. brownii (C4-like) and F. trinervia (C4) the PEPC amino acid sequence of F. brownii was more similar to the C3 and C3-C4 ppM PEPCs than to the C4 PEPC. Similarly, the C3, C3-C4 and C4-like ppM PEPCs showed almost identical PEP saturation kinetics when activated by glucose-6-phosphate (Km-PEP: 17-20 μM) while the Km-PEP for the C4 PEPC was determined to be 53 μM. However, without activation the ppc A PEPCs of F. pubescens and F. brownii displayed C3-C4 intermediate values. A similar picture was obtained when the malate sensitivities were compared. In the non-activated state the F. trinervia (C4) enzyme was 10 times more tolerant to malate than the F. pringlei counterpart. The ppcA enzymes of F. pubescens (C3-C4) and F. brownii (C4-like) displayed intermediate values. In contrast, the inclusion of 5 mM glucose-6-phosphate in the reaction mixture changed the order totally. Interestingly, the activation rendered the C4 enzyme about 50% less tolerant to malate than the C3 PEPC. The activation had a positive effect on malate tolerance of the F. pubescens (C3-C4) PEPC while the ppcA PEPC of F. brownii (C4-like) was almost unaffected. [ABSTRACT FROM AUTHOR]

Marine and terrestrial photosynthetic and chemoautotrophic microorganisms assimilate considerable amounts of carbon dioxide. Like green plastids, the predominant means by which this process occurs is via the Calvin-Benson-Bassham reductive pentose phosphate pathway, where ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco) plays a paramount role. Recent findings indicate that this enzyme is subject to diverse means of control, including specific and elaborate means to guarantee its high rate and extent of synthesis. In addition, powerful and specific means to regulate Rubisco activity is a characteristic feature of many microbial systems. In many respects, the diverse properties of microbial Rubisco enzymes suggest interesting strategies to elucidate the molecular basis of CO2/O2 specificity, the ‘holy grail’ of Rubisco biochemistry. These systems thus provide, as the title suggests, ‘different perspectives’ to this fundamental problem. These include vast possibilities for imaginative biological selection using metabolically versatile organisms with well-defined genetic transfer capabilities to solve important issues of Rubisco specificity and molecular control. This review considers the major issues of Rubisco biochemistry and regulation in photosynthetic microoganisms including proteobacteria, cyanobacteria, marine nongreen algae, as well as other interesting prokaryotic and eukaryotic microbial systems recently shown to possess this enzyme. [ABSTRACT FROM AUTHOR]

Two patients presented in early childhood with (i) alopecia, skin rashs, and dermatitis, (ii) severe hypotonia, ataxia and motor retardation, (iii) frequent episodes of ketoacidosis with hyperlactacidemia. Propionic and methylcrotonic aciduria only appeared on high protein diet. Mitochondrial biotin-dependent carboxylase activities were decreased in the liver and leukocytes, but were similar to control values in fibroblasts cultured in a biotin-free medium. In addition, the plasma biotin was found to be significantly lower than in control subjects. These disorders responded to biotin administration, pointing to biotin-dependent multiple carboxylase deficiencies (MCD). Our report stresses the polymorphism of MCD and suggests that MCD could be of two types: impaired vitamin metabolism (absorption, plasma transport), might result in low plasma biotin with generalized MCD involving acetyl CoA carboxylase. Defective mitochondrial holocarboxylase synthetase might lead to a pure mitochondrial MCD, with fibroblastic deficiency and presumably normal biotin metabolism. [ABSTRACT FROM AUTHOR]

The ac-20 mutant strain of the unicellular green alga, Chlamydomonas reinhardii, lacks both chloroplast ribosomes and ribulose bisphosphate carboxylase activity when grown on organic medium. Under these conditions, the cells do not posses pools of either the large or small subunit of this enzyme. When transferred to inorganic medium, the carboxylase activity recovers. During this recovery, de novo synthesis of both subunits occurs. Synthesis of both subunits is inhibited by chloramphenicol even when possible free subunit pools rather than just the subunits incorporated into whole enzyme are examined. [ABSTRACT FROM AUTHOR]

We have studied the problem whether the phytochrome-mediated accumulation of ribulosebisphosphate carboxylase ('carboxylase'; EC 4.1.1.39) in the cotyledons of sinapis alba L. is related to size, ultrastructure, or organization of the plastid compartment. We have shown that under different light conditions (e.g. continuous far-red light, continuous white light) which lead to conspicuously different plastids the time course of the enzyme levels remains precisely the same. It is concluded that the onset and the rate of carboxylase accumulation is not related to the organizational state of the plastid compartment as discernible under the electron microscope. [ABSTRACT FROM AUTHOR]

An acyl-coenzyme A carboxylase that carboxylates acetyl-CoA, butyryl-CoA, propionyl-CoA, and succinyl-CoA was purified from Myxococcus xanthus. Since the enzyme showed maximal rates of carboxylation with propionyl-CoA, the enzyme is thought to be propionyl-CoA carboxylase. The apparent K m values for acetyl-CoA, butyryl-CoA, propionyl-CoA, and succinyl-CoA were found to be 0.2, 0.2, 0.03, and 1.0 mM, respectively. The native enzyme has a molecular mass of 605–615 kDa and is composed of nonidentical subunits (α and β) with molecular masses of 53 and 56 kDa, respectively. The enzyme showed maximal activity at pH 7.0–7.5 and at 25–30°C, and was affected by variation in concentrations of ATP and Mg2+. During development of M. xanthus, the propionyl-CoA carboxylase activity increased gradually, with maximum activity observed during the sporulation stage. Previous work has shown that a propionyl-CoA-carboxylase-deficient mutant of M. xanthus reduces levels of long-chain fatty acids. These results suggest that the propionyl-CoA carboxylase is also responsible for the carboxylation of acetyl-CoA to malonyl-CoA used for the synthesis of long-chain fatty acids during development. [ABSTRACT FROM AUTHOR]

Short-chain aliphatic epoxides and ketones are two classes of toxic organic compounds formed biogenically and anthropogenically. In spite of their toxicity, these compounds are utilized as primary carbon and energy sources or are generated as intermediate metabolites in the metabolism of other compounds (e.g., alkenes, alkanes, and secondary alcohols) by a number of diverse bacteria. One bacterium capable of using both classes of compounds is the gram-negative aerobe Xanthobacter strain Py2. Studies of epoxide and ketone (acetone) metabolism by Xanthobacter strain Py2 have revealed a central role for CO2 in these processes. Both classes of compounds are metabolized by carboxylation reactions that produce β-keto acids as products. The epoxide- and ketone-converting enzymes are distinct carboxylases with molecular properties and cofactor requirements unprecedented for other carboxylases. Epoxide carboxylase is a four-component multienzyme complex that requires NADPH and NAD+ as cofactors. In the course of epoxide carboxylation, a transhydrogenation reaction occurs wherein NADPH undergoes oxidation and NAD+ undergoes reduction. Acetone carboxylase is a multimeric (three-subunit) ATP-dependent enzyme that forms AMP and inorganic phosphate as ATP hydrolysis products in the course of acetone carboxylation. Recent studies have demonstrated that acetone metabolism in diverse anaerobic bacteria (sulfate reducers, denitrifiers, phototrophs, and fermenters) also proceeds by carboxylation reactions. ATP-dependent acetone carboxylase activity has been demonstrated in cell-free extracts of the anaerobic acetone-utilizers Rhodobacter capsulatus, Rhodomicrobium vannielii, and Thiosphaera pantotropha. These studies have identified new roles for CO2 as a cosubstrate in the metabolism of two classes of important xenobiotic compounds. In addition, two new classes of carboxylases have been identified, the investigation of which promises to reveal new insights into biological strategies for the fixation of CO2 to organic substrates. [ABSTRACT FROM AUTHOR]

Following the analysis of transposon Tn 5432-induced mutants of Corynebacterium glutamicum ATCC 13032, a gene encoding a protein with a biotin-binding motif was cloned. The DNA sequence of this gene revealed an open reading frame encoding 591 amino acids with a calculated mol. mass of 63.4 kDa. The protein is composed of two domains, an N-terminal biotin carboxylase and a C-terminal biotin-carboxyl-carrier protein, that are highly similar to corresponding subunits from prokaryotic and eukaryotic biotin enzymes. Over 70% identity was found to a protein from Mycobacterium leprae proposed to be part of an acyl-CoA carboxylase. Since it was not possible to inactivate the C. glutamicum gene, the gene most likely encodes a subunit of the essential acetyl-CoA carboxylase, which catalyzes the committed step in fatty acid synthesis. [ABSTRACT FROM AUTHOR]

Sequence studies of the gene for the large subunit of ribulose-1,5-bisphosphate carboxylase from maize indicate the presence of two translation start sites, 18 bp apart. Each site is preceded by a suitable ribosome binding region. By using a simplified E. coli-based in vitro system which measures fromation of the first dipeptide of the gene product, we have determined that only the second methionine initiates translation. [ABSTRACT FROM AUTHOR]