Anaerobic Expression ofEscherichia coliSuccinate Dehydrogenase: Functional Replacement of Fumarate Reductase in the Respiratory Chain during Anaerobic Growth

Abstract

Succinate-ubiquinone oxidoreductase (SQR) from Escherichia coli is expressed maximally during aerobic growth, when it catalyzes the oxidation of succinate to fumarate in the tricarboxylic acid cycle and reduces ubiquinone in the membrane. The enzyme is similar in structure and function to fumarate reductase (menaquinol-fumarate oxidoreductase [QFR]), which participates in anaerobic respiration by E. coli. Fumarate reductase, which is proficient in succinate oxidation, is able to functionally replace SQR in aerobic respiration when conditions are used to allow the expression of the frdABCD operon aerobically. SQR has not previously been shown to be capable of supporting anaerobic growth ofE. coli because expression of the enzyme complex is largely repressed by anaerobic conditions. In order to obtain expression of SQR anaerobically, plasmids which utilize the PFRD promoter of the frdABCD operon fused to the sdhCDAB genes to drive expression were constructed. It was found that, under anaerobic growth conditions where fumarate is utilized as the terminal electron acceptor, SQR would function to support anaerobic growth ofE. coli. The levels of amplification of SQR and QFR were similar under anaerobic growth conditions. The catalytic properties of SQR isolated from anaerobically grown cells were measured and found to be identical to those of enzyme produced aerobically. The anaerobic expression of SQR gave a greater yield of enzyme complex than was found in the membrane from aerobically grown cells under the conditions tested. In addition, it was found that anaerobic expression of SQR could saturate the capacity of the membrane for incorporation of enzyme complex. As has been seen with the amplified QFR complex, E. coli accommodates the excess SQR produced by increasing the amount of membrane. The excess membrane was found in tubular structures that could be seen in thin-section electron micrographs.

Central pathway engineering for enhanced succinate biosynthesis from acetate in Escherichia coli

“…In E. coli, it contains two membrane-bound flavoproteins that catalyze the interconversion of fumarate and succinate, succinate dehydrogenase (SDH), and fumarate reductase (FRD) (Cecchini, Schröder, Gunsalus, & Maklashina, 2002). Previous reports already indicated that the FRD and SDH can be interchanged under optimal conditions (Guest, 1981;Maklashina, Berthold, & Cecchini, 1998).…”

Progress in understanding structure–function relationships in respiratory chain complex II

“…On the other hand, fumarate reduction by SQR shuts down' (70^90%) under conditions of high reducing potential (`tunnel diode e¡ect') [6]. Thus, SQRs appear ill-suited as physiological fumarate reducers, in accord with the observation that E. coli grows much less e¤ciently using SQR instead of QFR for anaerobic growth [7]. QFRs characteristically maintain catalytic e¤ciency under highly reducing conditions [6].…”

A thermodynamic atlas of carbon redox chemical space

“…The recently elucidated signalling functions 32,33 . Interestingly, succinate accumulation occurs in a number of different organisms, including bacteria such as Escherichia coli 34 , Mycobacterium tuberculosis 35 , as well as several bacterial members of the human gut microbiome [36][37][38][39] and the bovine rumen 40,41 ; fungi such as the yeast Saccharomyces cerevisiae 42 and members of the genus Penicillium 43 ; green algae 44 ; parasitic helminths 45 ; the sleeping sickness-causing parasite Trypanosoma brucei 46 ; marine invertebrates 47 ; and humans [48][49][50][51] . More specifically, our observations are consistent with the behavior of the TCA cycle under anaerobiosis and hypoxia [52][53][54][55] .…”