Mutants of Methylobacterium extorquens and Paracoccus denitrificans deficient in c‐type cytochrome biogenesis synthesise the methylamine‐dehydrogenase polypeptides but cannot assemble the tryptophan‐tryptophylquinone group

Abstract

Five mutants of Methylobacterium extorquens and four mutants of Paracoccus denitrificans that have a general defect in c-type cytochrome synthesis also failed to assemble an active methylamine dehydrogenase. In all cases methanol dehydrogenase, another periplasmic enzyme, was fully active. All nine mutant strains accumulated both the heavy and light subunits of methylamine dehydrogenase to essentially wild-type levels. In all nine mutants, the heavy-subunit and light-subunit polypeptides were proteolytically processed, suggesting that translocation to the periplasm had occurred; in the case of the f? denitrificans mutants, a periplasmic location for the heavy and light subunits was confirmed experimentally. While specific quinone staining of the methylamine dehydrogenase light subunit in wild-type M. extorquens and f? denitrificans strains could readily be demonstrated, the light subunit polypeptides accumulated by the mutants did not quinone stain, indicating that the methylamine dehydrogenase prosthetic group, tryptophan tryptophylquinone, is not assembled in the absence of functional c-type cytochromes.Methylamine dehydrogenase, catalysing the oxidation of methylamine to formaldehyde and ammonia, has been identified in several groups of Gram-negative bacteria, the facultative (e.g. Methylobacterium extorquens) and restricted facultative (e.g. Methylophilus spp.) methylotrophs and the facultative autotrophs (e.g. Paracoccus denitrificans) (Eady and Large, 1968;Haywood et al., 1982;Husain and Davidson, 1987). The enzymes have an a& subunit structure with a heavy (H) subunit of 40-45 kDa and light (L) subunit of 14-15 kDa; in all known cases the enzyme is periplasmic (Burton et al., 1983;Husain and Davidson, 1987;Ubbink et al., 1991;Chistoserdov et al., 1992). The methylamine dehydrogenase prosthetic group, recently shown to be a unique quinonoid moiety termed tryptophan tryptophylquinone (TTQ), is formed from two tryptophan residues (positions 55 and 106 in the M . extorquens enzyme; Ishii et al., 1983, Chistoserdov et al., 1990 in the L-subunit polypeptide chain (McIntire et al., 1991). The tryptophan residue that occurs nearer the N-terminal in the amino acid sequence (1.e.

Structure of a quinohemoprotein amine dehydrogenase with an uncommon redox cofactor and highly unusual crosslinking

“…It has been shown that the TPQ cofactor in CAO (35) and the Cys-Tyr crosslinked cofactor in galactose oxidase (36) are both generated by the copperdependent autocatalytic process of the precursor proteins. On the other hand, the biosynthesis of TTQ in MADH appears to be catalyzed by an enzymatic process, because the absence of one of the genes (mauG) of the 11 genes in the mau operon prevents the maturation of TQQ (37). The biosynthetic process for CTQ in the ␥ subunit of QHNDH may involve an enzyme-mediated process as well.…”

The Cytochrome c Maturation Locus of Legionella pneumophila Promotes Iron Assimilation and Intracellular Infection and Contains a Strain-Specific Insertion Sequence Element

“…That ccm mutations have pleiotropic effects is becoming increasingly apparent. In the extracellular bacterial systems, ccm has been implicated in manganese oxidation (17,24), gluconate and 2-ketogluconate oxidation (70), methylamine oxidation (64), nicotinic acid hydroxylation (49), copper resistance (86), and the cis-trans isomerization of unsaturated fatty acids (42). Thus, future characterization of our infectivity mutants should provide new insights into ccm and cytochrome c function in particular and bacterial physiology and intracellular infection in general.…”

The Covalent Structure of the Small Subunit fromPseudomonas putida Amine Dehydrogenase Reveals the Presence of Three Novel Types of Internal Cross-linkages, All Involving Cysteine in a Thioether Bond

“…Examples of an autocatalytic system are the biogenesis of the topaquinone cofactor in copper amine oxidases (15) and of the Cys-Tyr cofactor in galactose oxidase (16), in the latter case with the formation of a thioether linkage. An example of an enzymatic process is the biosynthesis of tryptophan tryptophylquinone in methylamine dehydrogenase, where the absence of one of the genes (mauG) of the 11 genes in the mau operon appears to prevent the maturation of tryptophan tryptophylquinone (17,18).…”