Isolation of a fourth cysteinyl-containing peptide of the alpha-subunit of the F1 ATPase from Escherichia coli necessitates revision of the DNA sequence

The rapid determination of cysteinyl residues by Creighton's method [(1980) Nature 284, 487-489] led to the discovery of a discrepancy between protein and DNA sequence data in the alpha-subunit of the F1 ATPase from Escherichia coli [(1984) Arch. Biochem. Biophys. 229, 320-328]. We have isolated a cysteinyl-containing decapeptide from the alpha-subunit with a protein sequence (AGCAMGEYFR) which is only partially recognizable from DNA data. Re-sequencing of DNA in the region coding for the peptide has resulted in two corrections: insertion of a cytosine before position 715 and deletion of a thymine at position 731 of the uncA gene.     

Inhibition by cyanide of the respiratory chain oxidases of Escherichia coli

The kinetics of inhibition by KCN of NADH oxidation in respiratory particles from Escherichia coli could be related to the relative amounts of cytochromes d and o which were present. Particles which contained higher levels of cytochrome d relative to cytochrome o were less sensitive to inhibition by cyanide. When cyanide reacted with the respiratory particles, the absorption bands of reduced cytochrome d at 442 and 628 nm in the reduced plus cyanide minus reduced difference spectrum were eliminated, as also were the bands at 423, 428, and 555 nm of b- and/or c-type cytochromes.Cyanide appeared to react with the oxidized form of cytochrome d to eliminate its α-band absorption with a second-order rate constant of 0.011 m^?1 sec^?1 for the rate of formation of cyanocytochrome d in the absence of added substrate. Under turnover conditions using NADH as substrate, the rate constant was 0.58 m^?1 sec^?1. This value is close to that determined from cyanide inhibition of NADH oxidase activity. The magnitude of the second-order rate constant for the formation of cyanocytochrome d was directly related to the rate of electron flux through cytochrome d. It is suggested that an intermediate species formed during the normal oxidation-reduction cycle of cytochrome d reacts with cyanide.     

Loss of protection by nucleotides against proteolysis and thiol modification in the isolated alpha-subunit from F1 ATPase of Escherichia coli mutant uncA401.

Binding of nucleotides to the high-affinity site of the isolated alpha subunit of normal Escherichia coli F1 adenosine triphosphatase (ATPase) results in partial protection against digestion by trypsin [Senda, Kanazawa, Tsuchiya & Futai (1983) Arch. Biochem. Biophys. 220, 398-440]. In contrast, the isolated alpha subunit from the defective ATPase of the E. coli uncA401 mutant (strain AN120) is cleaved by trypsin to peptides of less than 8000 Da in the presence of ADP or ATP (2.5 microM-110 mM). The nucleotide-dependent accessibility of thiol groups of the isolated alpha subunit was also studied. Two out of four thiol groups of the alpha subunit from normal ATPase are labelled by fluorescent maleimides or iodoacetates, but in the presence of ADP or ATP (0.14-1.2 mM), reaction of thiol groups with these labels is almost absent. Mutant alpha subunit, however, is labelled by these reagents at all four thiol groups in the presence or absence of ADP or ATP (1 mM). These results suggest that the mutation in the ATPase of strain AN120 leads either to the loss of the high-affinity nucleotide-binding site or affects transmission of allosteric changes that occur on binding of nucleotide to the isolated alpha subunit.     

Molecular characterization of a heat-modifiable protein from the outer membrane of Escherichia coli.

Discrete fractions of nonhistone chromosomal proteins (NHCP) were obtained from rabbit liver chromatin by their dissociation in 5 m urea with increasing concentrations of NaCl. Three fractions were obtained: M₀, M₁, and M₃. We found that M₀ can modify the conformation of DNA/histone complexes as depicted from their induced increase in the ellipticity of DNA/histone from 5100 to 6900 degree-cm²/dmol. This effect was found to be reversible, while M₁ and M₃ effects, if any, were not measurable. These results suggest that M₀ primarily interacts with the chromatin subunit.     

Predictive habitat modelling to estimate petrel breeding colony sizes: Sooty shearwaters (Puffinus griseus) and mottled petrels (Pterodroma inexpectata) on Whenua Hou Island

Abstract

Between 2001 and 2006, we systematically sampled the entire coast of Whenua Hou, a rugged offshore island in southern New Zealand, to estimate the population densities of sooty shearwaters (Puffinus griseus) and mottled petrels (Pterodroma inexpectata) by counting the entrances of breeding burrows. A two‐step regression modelling process using binomial errors was used to predict the presence of a colony, and a normal general linear model was used to predict the density of entrances within colonies. Aerial photography, GIS and a Digital Elevation Model were used to extract relevant habitat and location variables, and a combination of both regression models was used to predict the density of breeding burrows within each 5.32 m² pixel on the island. This complex GIS and habitat prediction modelling approach gave population estimates very similar to a more traditional simple area extrapolation method and gave no improvement in precision. However, correction for the slope of the land increased our simple area estimates of population size by 11%. We estimate populations of sooty shearwater and mottled petrel breeding pairs at 173 000 (162 000–190 000) and 160 000 (123 000–197 000) respectively. Based on this number of breeding pairs, we calculate that Whenua Hou supports a total population of 868 000 (554 000–1 270 000) sooty shearwaters. Our estimate of the total mottled petrel population 202 000 pairs (162 000–242 000) is comparable with the only published estimate, but could be an underestimate because mottled petrels are sometimes found in large burrows. More research for robust estimation of population trends is needed to assess the conservation status of mottled petrels.