Rubredoxin reductase from Alcanivorax borkumensis: Expression and characterization

Abstract

Oil pollution is an environmental problem of increasing importance. Alcanivorax borkumensis, with a high potential for biotechnological applications, is a key marine hydrocarbonoclastic bacterium and plays a critical role in the bioremediation of oil-polluted marine systems. In oil degrading bacteria, the first step of alkane degradation is catalyzed by a monooxygenase. The reducing electrons are tunneled from NAD(P)H via rubredoxin, one of the most primitive metalloproteins, to the hydroxylase. Rubredoxin reductase is a flavoprotein catalyzing the reduction of rubredoxin. There are two rubredoxin genes, alkG and rubA, in A. borkumensis genome. In this work, the genes encoding rubredoxin reductase (ABO_0162, rubB) and AlkG(ABO_2708, alkG) were cloned and functionally overexpressed in E. coli. Our results demonstrate that RubB could reduce AlkG, therefore compensating for the absence of AlkT, also a rubredoxin reductase, missing in A. borkumensis SK2 genome. These results will increase our knowledge concerning biological alkane degradation and will lead us to design more efficient biotransformation and bioremediation systems.

Differential protein expression during growth on linear versus branched alkanes in the obligate marine hydrocarbon‐degrading bacterium Alcanivorax borkumensis SK2T

“…Bacteria degrading linear alkanes typically contain a non‐heme alkane oxygenase system including a membrane bound alkane monooxygenase, a rubredoxin and a rubredoxin reductase and/or a heme‐thiolate cytochrome system including a soluble cytochrome P450, a ferredoxin and ferredoxin reductase (Teimoori et al, ; Nie et al, ). It is rather common to find bacteria that contain more than one alkane oxidation system as each have different substrate ranges or different induction patterns.…”

“…Rubredoxin reductases oxidizes NAD(P)H to transfer electrons to rubredoxin reducing it allowing electron transfer to the monooxygenase. There is one rubredoxin reductase gene in the A. borkumensis genome, RubB (ABO_0162), which has been shown to reduce AlkG when both proteins were cloned and functionally overexpressed in E. coli (Teimoori et al, ). This activity compensates for a lack of AlkT, also a rubredoxin reductase, missing in the A. borkumensis genome (Schneiker et al, ).…”

Identification of Electrode Respiring, Hydrocarbonoclastic Bacterial Strain Stenotrophomonas maltophilia MK2 Highlights the Untapped Potential for Environmental Bioremediation

“…Such a successional oxidation process is activated by a family of integral membrane proteins called alkane hydroxylase enzyme system, and this was first studied in Pseudomonas putida GPo1. This is of particular interest being a three component biocatalyst and composed of alkane monooxygenase ( alk B group), dinuclear iron rubredoxins ( rub A, rub B) and mononuclear rubredoxin reductase ( rub R) (Rojo, 2009; Teimoori et al, 2011). These genes encode the alkane hydroxylase ( alk ) system in the enzyme complex which activates the terminal carbon atoms in the chain of hydrocarbons.…”

Biochemical characterization of two recombinant ferredoxin reductases from Alcanivorax borkumensis SK2

“…The adsorbed proteins were then eluted with a linear gradient of sodium chloride in the range of 0–0.5 M in the equilibration buffer. Two milliliters of fractions was collected at a flow rate of 20 mL/H and analyzed for the protein content .…”