Purification and Biochemical Characterization of the F₁-ATPase fromAcidithiobacillus ferrooxidansNASF-1 and Analysis of theatpOperon

Abstract

ATPase was purified 51-fold from a chemoautotrophic, obligately acidophilic iron-oxidizing bacterium, Acidithiobacillus ferrooxidans NASF-1. The purified ATPase showed the typical subunit pattern of the F 1 -ATPase on a polyacrylamide gel containing sodium dodecyl sulfate, with 5 subunits of apparent molecular masses of 55, 50, 33, 20, and 18 kDa. The enzyme hydrolyzed ATP, GTP, and ITP, but neither UTP nor ADP. The K m value for ATP was 1.8 mM. ATPase activity was optimum at pH 8.5 at 45 C, and was activated by sulfite. Azide strongly inhibited the enzyme activity, whereas the enzyme was relatively resistant to vanadate, nitrate, and N,N 0 -dicyclohexylcarbodiimide. The genes encoding the subunits for the F 1 F O -ATPase from A. ferrooxidans NASF-1 were cloned as three overlapping fragments by PCR cloning and sequenced. The molecular masses of the , , , , and " subunits of the F 1 portion were deduced from the amino acid sequences to be 55.5, 50.5, 33.1, 19.2, and 15.1 kDa, respectively.

Modelling of the dynamic polarizability of macromolecules for single-molecule optical biosensing

“…The presence of light can also cause aggregation of molecules, and alter the function of molecular machines. For instance, aggregation has been observed to occur at light intensities of 1 for -chymotrypsin molecules attached to gold nanoparticles in solution 67 , while the function of ATPase is known to degrade at temperatures above C 68 , corresponding to an intensity of about for a plasmonic trap sensor.…”

Molecular aspects of bacterial pH sensing and homeostasis

“…An acidophile F 1 F o -ATP synthase with the typical eight subunits has a pH optimum of 8.5 for its hydrolytic activity in assays of the membrane-associated enzyme; the proton-translocating a- and c- subunits of the enzyme have some deviations from neutralophile synthases but whether these are adaptive to the unusual PMF pattern is not yet known 116 . The acidophile PMF pattern results in sensitivity of these bacteria to organic acids, because of the large ΔpH, and to toxic anions, because of the inside-positive Δψ 9 .…”

Oxidative phosphorylation in a thermophilic, facultative chemoautotroph, Hydrogenophilus thermoluteolus, living prevalently in geothermal niches

“…A), which was 30% of that with MgCl 2 . Ca 2+ , in addition to Mg 2+ , is also similarly effective for various F‐type ATPases from other bacteria (Wakai et al ., ; Ferguson et al ., ). At the optimal pH of 9.0, the ATPase activity of the purified H. thermoluteolus F1‐ATPase increased as the temperature increased from 10 to 65°C, and disappeared at 80°C (Fig.…”