Molecular cloning and primary structure of the human blood group RhD polypeptide.

The RH (rhesus) blood group locus from RhD-positive donors is composed of two homologous structural genes, one of which encodes the Cc and Ee polypeptides, whereas the other, which is missing in the RhD-negative condition, encodes the D protein that carries the major antigen of the RH system. Recently, different splicing isoforms transcribed from the CcEe gene were isolated. We report now the characterization of two other Rh clones, RhII and RhXIII, generated by alternative choices for poly(A) addition sites that were identified as the RhD gene transcripts. That these cDNAs represented the RhD messenger and that the previously described Rh clones were derived from the CcEe gene was demonstrated by amplification of RhII/XIII sequences only from D-positive genomes and by cloning and sequencing of D- and CcEe-specific gene fragments. The predicted translation product of the RhD mRNA is a 417-amino acid protein (M(r) = 45,500) that exhibited a similar membrane organization with 13 bilayer-spanning domains compared with the polypeptide encoded by the CcEe gene. The D and Cc/Ee polypeptides differ by 36 amino acid substitutions (8.4% divergence), but the NH2- and COOH-terminal regions of the two proteins are well conserved. Similarly, five of the six cysteine residues of the Cc/Ee proteins were conserved in the D protein, including the unique exofacial cysteine, which is critical for antigenic reactivity. The sequence homology between the Cc/Ee and D proteins supports the concept that the genes encoding these polypeptides have evolved by duplication of a common ancestor gene.

[Study of the NADH and NADPH-ferredoxin oxidoreductase activities in Clostridium acetobutylicum]. / Etude des activités NADH et NADPH-ferrédoxine oxydoréductasiques chez Clostridium acetobutylicum

The NADH and NADPH-ferredoxin oxidoreductase have been studied in Clostridium acetobutylicum. Acetyl-CoA is an obligatory activator of NADH-ferredoxin reductase activity and NADH a competitive inhibitor of ferredoxin-NAD+ reductase activity. These regulations are the same when C. acetoburylicum moves from 'butylic-type metabolism' to 'butyric-type metabolism'; this demonstrates that NADH-ferredoxin oxidoreductase cna, through its reversible action, meet the very different cell needs imposed by these two types of culture. The physiological function of the clostridial NADPH-ferredoxin oxidoreductase was anabolic as it has been with other clostridia.

Regulation of the NADH and NADPH-ferredoxin oxidoreductases in clostridia of the butyric group.

NADH and NADPH-ferredoxin oxidoreductases have been studied in Clostridium acetobutylicum, Cl. tyrobutyricum and Cl. pasteurianum. The study of the distribution and regulation of these enzymatic activities in well-defined culture conditions, reveals that the essential function of NADPH-ferredoxin oxidoreductase is to produce NADPH, while NADH-ferredoxin oxidoreductase can, depending on cellular conditions, produce or oxidize NADH. When these Clostridia use glycolysis, regulation of the NADH-ferredoxin oxidoreductase by acetyl-CoA (obligatory activator of NADH-ferroxin reductase activity) and by NADH (competitive inhibitor of ferredoxin-NAD+ reductase activity) allow the enzymes to function correlatively with glyceraldehyde-3-phosphate dehydrogenase and thus control the levels of NAD+ and NADH in the cell. In Cl. tyrobutyricum and Cl. pasteurianum, the ferredoxin-NADP+ reductase activities are regulated by NAD+ and NADH in accordance with the intracellular concentrations of these coenzymes. In Cl. tyrobutyricum growing on pyruvate/acetate, NADH and NADPH-ferredoxin reductase activities cannot be detected; only the ferredoxin-NAD+ and ferredoxin-NADP+ reductase activities are found. In this Clostridium, regulation of the ferredoxin-NADP+ reductase activity is the same whether it is grown on glucose or pyruvate. Contrary to this, the ferredoxin-NAD+ reductase activity undergoes a drastic change, since NADH no longer controls the enzymatic activity. In this case regulation is no longer necessary, since glyceraldehyde-3-phosphate dehydrogenase does not function.

[The spore germination of "Clostridium tyrobutyricum" III.--An hypothesis on the mechanism of initiation (author's transl)]. / La germination de la spore de Clostridium tyrobutyricum III.--Hypothèse sur le mécanisme de la phase initiale

Spores of C. tyrobutyricum do not contain 3-phosphoglyceric acid (PGA) but a polysaccharide which could replace PGA as an energy source during germination. The absence of PGA, which is an inhibitor of phosphotransacetylase, confirms the role of the acetyl-CoA synthesizing system in the germination initiated by acetate. Spore extracts of C. tyrobutyricum, as extracts of vegetative cells, were found to contain a ferredoxin and exhibited a NADH-ferredoxin oxydase activity which required the presence of an acetyl-CoA regenerating system, suggesting that this enzyme is also involved in germination. From this results, an hypothesis on the role of initiators (acetate and NH4+) in the mechanism of initiation of spore germination in C. tyrobutyricum is proposed. Acetate would have an effect on the utilisation of the endogenous polysaccharide and on glucose catabolism, and therefore, would be an effector for the production of the energy required particularly to transport cations into the spore.