Molybdate‐Uptake Genes and Molybdopterin‐Biosynthesis Genes on a Bacterial Plasmid

Abstract

A gene cluster consisting of homologs to Escherichia coli moaA, moeA, moaC and moaE, which encode enzymes involved in the biosynthesis of molybdopterin cofactor (MoCo), and to modA, modB and modC, which encode a high-affinity molybdate transporter, were identified on pAO1 of Arthrobacter nicotinovorans near genes of molybdopterin-dependent enzymes involved in nicotine degradation. This gene arrangement suggests a coordinated expression of the MoCo-dependent and the MoCo-biosynthesis genes and shows that catabolic plasmids may carry the transport and biosynthetic machinery for the synthesis of the cofactors needed for the functioning of the enzymes they encode. pAO1 MoeA functionally complemented E. coli moeA mutants. The overexpressed and purified protein, of molecular mass 44 500 Da, associated into high-molecular-mass complexes and spontaneously formed gels at concentrations above 1 mg/ml. Transmission electron microscopy and atomic force microscopy revealed that MoeA forms fibrilar structures. In the presence of Mg 2ϩ MoeA exhibited ATPase activity (0.020 pmol ATP · pmol protein Ϫ1 · min Ϫ1 ). ATP, ADP or AMP induced the disassembly of the MoeA fibers into aggregates. pAO1 MoeA shows 39% identity to the C-terminal domain of the rat neuroprotein gephyrin. Like gephyrin it binds to neurotubulin, but binds with preference to tubulin dimers.

The GDP-GTP Exchange Factor Collybistin: An Essential Determinant of Neuronal Gephyrin Clustering

“…Furthermore, our results with deletion constructs indicated that the binding site for GlyR ␤ must reside entirely within the MoeA homology domain (amino acids 323-736). If the gephyrin MoeA homology domain forms an antiparallel dimer, like E. coli MoeA (Menéndez et al, 1997;Schrag et al, 2001;Xiang et al, 2001), our results strongly suggest that two equivalent GlyR ␤ binding sites would be present at the MoeA-MoeA homology domain interface. Whether these two equivalent sites would bind a single GlyR heteromer, which is thought to contain two ␤ subunits (stoichiometry ␣1 3 ␤ 2 ) or two distinct receptor pentamers, is presently unclear.…”

The enzymes of microbial nicotine metabolism

“…Comparison of the pAO1 sequence with that of xanthine oxidoreductase identified ndhs , ndhm , and ndhl (initially designated ndhABC ) as coding for three proteins: a 14.9 kDa subunit containing an iron–sulfur cluster, a 30 kDa subunit with an FAD binding site, and an 87.7 kDa subunit containing the molybdopterin site, respectively [9–10]. Consistent with this identification, expression of the active enzyme required molybdopterin [9], and pAO1 contains a number of genes that have been identified as coding for proteins involved in uptake of molybdenum and biosynthesis of the molybdopterin cofactor [11]. The mechanism of Scheme 2 can be written for nicotine dehydrogenase by analogy to the mechanism of xanthine oxidoreductase [8].…”

Characterization of the ModABC Molybdate Transport System of Pseudomonas putida in Nicotine Degradation

“…Mo uptake occurred during nicotine biodegradation by gram-positive bacteria (Freudenberg et al, 1988; Menendez et al, 1997). The nicotine utilization of Arthrobacter oxidans P-34 (DSM419) required a supplementation with molybdate to the growth medium, and nicotine dehydrogenase (NDH) was identified as a molybdo-iron-sulfur-flavoprotein (Freudenberg et al, 1988).…”