Form of inorganic carbon involved as a product and as an inhibitor of c(4) Acid decarboxylases operating in c(4) photosynthesis

These studies demonstrated that CO₂ rather than HCO₃⁻ is the inorganic carbon metabolite produced by the C₄ acid decarboxylases involved in C₄ photosynthesis (chloroplast located NADP malic enzyme, mitochondrial NAD malic enzyme, and cytosolic phosphoenolpyruvate [PEP] carboxykinase). The effect of varying CO₂ or HCO₃⁻ as a substrate for the carboxylation reaction catalyzed by these enzymes or as inhibitors of the decarboxylation reaction was also determined. The K_mCO₂ was 1.1 millimolar for NADP malic enzyme and 2.5 millimolar for PEP carboxykinase. For these two enzymes the velocity in the carboxylating direction was substantially less than for the decarboxylating direction even with CO₂ concentrations at the upper end of the range of expected cellular levels. Activity of NAD malic enzyme in the carboxylating direction was undetectable. The decarboxylation reaction of all three enzymes was inhibited by added HCO₃⁻. For NADP malic enzyme CO₂ was shown to be the inhibitory species but PEP carboxykinase and NAD malic enzyme were apparently inhibited about equally by CO₂ and HCO₃⁻.     

Purification and steady-state kinetic characterization of human liver beta 3 beta 3 alcohol dehydrogenase

Human liver alcohol dehydrogenase catalyzes the NAD+-dependent oxidation of alcohols. Isoenzymes are produced in liver by five different genes, two of which are polymorphic. We have studied the three beta beta isoenzymes produced at ADH2 because they exhibit very different kinetic properties and they appear with different frequencies in different racial populations. The beta 3 beta 3 isoenzyme which appears in 25% of black Americans was purified to homogeneity, and conditions were found to stabilize this labile isoenzyme. The comparison of substrate specificity among beta beta isoenzymes for primary straight-chain alcohols indicates that there is a positive correlation between Vmax/KM and the log octanol/water partition coefficient for alcohols with beta 2 beta 2 and beta 3 beta 3 but not with beta 1 beta 1. Methyl substitutions at C1 or C2 of these alcohols reduce the catalytic efficiency with all three isoenzymes. The KM and Ki values of beta 3 beta 3 for NAD+ and NADH are substantially higher than values for beta 1 beta 1 or beta 2 beta 2. The Vmax of beta 3 beta 3 ethanol oxidation is 90 times that of beta 1 beta 1. Sequencing of the beta 3 subunit and gene indicates that the polymorphism results from a single amino acid exchange of Cys-369 in beta 3 for Arg-369 in beta 1 and beta 2 [Burnell et al. (1987) Biochem. Biophys. Res. Commun. 146, 1227-1233]. In horse alcohol dehydrogenase and beta 1 beta 1, the guanidino group of Arg-369 is thought to stabilize the NAD(H)-enzyme complex by bonding to one of the pyrophosphate oxygens.(ABSTRACT TRUNCATED AT 250 WORDS)     

Spinach chloroplastic carbonic anhydrase: nucleotide sequence analysis of cDNA

We have determined the nucleotide sequence of a cDNA encoding spinach (Spinacia oleracea) chloroplastic carbonic anhydrase (CA). The open reading frame encodes a protein consisting of a transit peptide and a mature CA protein with a predicted mass of 24, 116 daltons. This represents the first report of a nucleotide sequence of a plant CA.     

A linkage study in seven breast cancer families.

Seven breast cancer families are examined for evidence of linkage to a site in the region of 17q12-q21, by using five markers. The families constitute a subset of a larger series of familial breast cancer; the seven families were selected because constitutional DNA was available on informative members, either from clinical samples or extracted from paraffin blocks. Two-point lod scores are reported. The maximum lod score, 0.8824, is obtained with marker NM23 at theta = 0. This is clearly not significant in itself; however, when taken in context with evidence from existing reports, it provides support for linkage to this region.     

The rDNA Internal Transcribed Spacer Region as a Taxonomic Marker for Nematodes

The ITS region from a wide taxonomic range of nematodes, including secernentean and adenophorean taxa, and free-living, entomopathogenic, and plant-parasitic species, was evaluated as a taxonomic marker. Size of the amplified product aided in the initial determination of group membership, and also suggested groups that may require taxonomic reevaluation. Congeneric species often displayed identically sized ITS regions, but genera such as Pratylenchus and Tylenchorhynchus had species with large differences in size. ITS heterogeneity in individuals and populations was identified in several nematode taxa. PCR-RFLP of ITS1 is advocated as a method of taxonomic analysis in genera such as Helicotylenchus that contain numerous species with few diagnostic morphological characteristics.     

Variations in the levels of asterone and asterogenol, two steroids from the saponins of the starfish, Asterias vulgaris (Verrill)

The levels of two steroids, asterone 1 and asterogenol 4, obtained by hydrolysis of the crude asterosaponin mixture from the starfish Asterias vulgaris, were highest in winter and spring, then the steroid levels fell to their annual minima in July, after the spawning period. Levels also varied geographically, but the ratio of these steroids remained approximately constant.     

Regulation of C4 photosynthesis: identification of a catalytically important histidine residue and its role in the regulation of pyruvate,Pi dikinase

These studies provide further information regarding the mechanism of the light/dark-mediated regulation of pyruvate,Pi dikinase in leaves. It is shown that a catalysis-linked phosphorylation of pyruvate,Pi dikinase can be demonstrated following incubation of the enzyme with [32P]phosphoenolpyruvate or [beta-32P]ATP plus Pi, that the enzyme-bound phosphate is located on a histidine residue, and that this phosphate is retained during ADP-mediated inactivation. Further evidence is provided that phosphorylation of this histidine is a prerequisite for ADP-mediated inactivation through phosphorylation of a threonine residue from the beta-phosphate of ADP. It is demonstrated that diethylpyrocarbonate (which forms a derivative with histidine residues) prevents [32P]phosphoenolpyruvate-dependent labeling (catalytic labeling) and [beta-32P]ADP-dependent labeling (inactivation labeling) of the enzyme. In addition, it is demonstrated that oxalate, an analog of pyruvate, competitively inhibits ADP-dependent inactivation with respect to ADP. The significance of these results is discussed with regard to the mechanism of regulation of pyruvate,Pi dikinase in vivo.     

Cysteinyl-tRNA Synthetase from Astragalus Species

l-Cysteinyl-tRNA synthetases (EC 6.1.1.16) from four Astragalus species were partially purified. The substrate specificities of the cysteinyl-tRNA synthetase from three selenium accumulator species (A. crotalariae, A bisulcatus, and A. racemosus) were compared with those from two nonaccumulator species (A. lentigenosus and Phaseolus aureus). All species had similar K(m) values for cysteine, selenocysteine, and alpha-aminobutyric acid except A. bisulcatus which failed to use selenocysteine as a substrate and which had a K(m) for cysteine four times greater than the K(m) values for other species.