Abstract
Based on a bioinformatics study, the protein MA4561 from the methanogenic archaeon Methanosarcina acetivorans was originally predicted to be a multidomain phytochrome-like photosensory kinase possibly binding open-chain tetrapyrroles. Although we were able to show that recombinantly produced and purified protein does not bind any known phytochrome chromophores, UV-visible spectroscopy revealed the presence of a heme tetrapyrrole cofactor. In contrast to many other known cytoplasmic heme-containing proteins, the heme was covalently attached via one vinyl side chain to cysteine 656 in the second GAF domain. This GAF domain by itself is sufficient for covalent attachment. Resonance Raman and magnetic circular dichroism data support a model of a six-coordinate heme species with additional features of a five-coordination structure. The heme cofactor is redox-active and able to coordinate various ligands like imidazole, dimethyl sulfide, and carbon monoxide depending on the redox state. Interestingly, the redox state of the heme cofactor has a substantial influence on autophosphorylation activity. Although reduced protein does not autophosphorylate, oxidized protein gives a strong autophosphorylation signal independent from bound external ligands. Based on its genomic localization, MA4561 is most likely a sensor kinase of a two-component system effecting regulation of the Mts system, a set of three homologous corrinoid/methyltransferase fusion protein isoforms involved in methyl sulfide metabolism. Consistent with this prediction, an M. acetivorans mutant devoid of MA4561 constitutively synthesized MtsF. On the basis of our results, we postulate a heme-based redox/dimethyl sulfide sensory function of MA4561 and propose to designate it MsmS (methyl sulfide methyltransferase-associated sensor).
Highlights
Multidomain sensory kinases are involved in numerous receptive processes in all kingdoms of life
The Putative Sensor Kinase MA4561—In silico analysis of MA4561 revealed the presence of two alternating N-terminal PAS and GAF domains fused to a C-terminal H_ATPase domain (Figs. 1 and 2A)
MA4561 is unable to bind any known chromophores of phytochrome photoreceptors. Based on this result and because it is unknown whether the methanogenic archaeon M. acetivorans is able to produce open-chain tetrapyrroles as their classical biosynthesis requires molecular oxygen, we rule out the possibility that this protein is a phytochrome
Summary
Multidomain sensory kinases are involved in numerous receptive processes in all kingdoms of life. Based on its genomic localization, MA4561 is most likely a sensor kinase of a two-component system effecting regulation of the Mts system, a set of three homologous corrinoid/methyltransferase fusion protein isoforms involved in methyl sulfide metabolism. Consistent with this prediction, an M. acetivorans mutant devoid of MA4561 constitutively synthesized MtsF. Upon signal recognition by the sensor kinase, a conserved histidine residue is autophosphorylated, and the phosphoryl group subsequently is transferred to a conserved aspartate residue in the response regulator input domain, resulting in activation of the output domain. As deletion of MA4561 led to deregulated synthesis of a dimethyl sulfide-dependent methyltransferase, we argue for a redox/dimethyl sulfide signaling function for MA4561
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