Literature summary for 1.14.13.25 extracted from

Kim, H.; An, S.; Park, Y.R.; Jang, H.; Yoo, H.; Park, S.H.; Lee, S.J.; Cho, U.S.
MMOD-induced structural changes of hydroxylase in soluble methane monooxygenase (2019), Sci. Adv., 5, eaax0059 .

Search for more literature for 1.14.13.25

Cloned(Commentary)

Cloned (Comment)	Organism
genes mmoB encoding MMOB and mmoC encoding MMOR, recombinant expression of subunits in Escherichia coli BL21(DE3), and gene mmoD encoding MMOD is recombinantly expressed as His-MBP-tagged protein in Escherichia coli strain Rosetta (DE3)	Methylosinus sporium

Crystallization (Commentary)

Crystallization (Comment)	Organism
purified recombinant MMOH-MMOD complex, hanging drop vapor diffusion method, mixing of 10 mg/ml subunits protein solutions containing 30 mM HEPES, pH 7.5, 100 mM NaCl, and 1 mM TCEP in a 1:2 molar ratio, and mixing with the crystallization solution containing with a mixture of 10% w/v PEG 8000, 20% v/vethylene glycol, 0.02 M 1,6-hexanediol, 0.02 M 1-butanol, 0.02 M 1,2-propanediol, 0.02 M 2-propanol, 0.02 M 1,4-butanediol, 0.02 M 1,3-propanediol, 0.89 M 1,3-butanediol, and 0.1 M MES/imidazole buffer, pH 6.5, one week at room temperature, X-ray diffraction structure determination and analysis at 2.6 A resolution, molecular replacement using Methylosinus trichosporium strain OB3b MMOH as the search model (PDB ID 1MHY), and modeling	Methylosinus sporium

Crystallization (Comment)

Organism

purified recombinant MMOH-MMOD complex, hanging drop vapor diffusion method, mixing of 10 mg/ml subunits protein solutions containing 30 mM HEPES, pH 7.5, 100 mM NaCl, and 1 mM TCEP in a 1:2 molar ratio, and mixing with the crystallization solution containing with a mixture of 10% w/v PEG 8000, 20% v/vethylene glycol, 0.02 M 1,6-hexanediol, 0.02 M 1-butanol, 0.02 M 1,2-propanediol, 0.02 M 2-propanol, 0.02 M 1,4-butanediol, 0.02 M 1,3-propanediol, 0.89 M 1,3-butanediol, and 0.1 M MES/imidazole buffer, pH 6.5, one week at room temperature, X-ray diffraction structure determination and analysis at 2.6 A resolution, molecular replacement using Methylosinus trichosporium strain OB3b MMOH as the search model (PDB ID 1MHY), and modeling

Methylosinus sporium

Inhibitors

Inhibitors	Comment	Organism	Structure
additional information	inhibitory role of subunit MMOD. MMOD binding leads to steric hindrance with the N-terminal region of MMOHbeta, which then triggers the detachment of MMOHbeta-NT from MMOHalpha	Methylosinus sporium

KM Value [mM]

KM Value [mM]	KM Value Maximum [mM]	Substrate	Comment	Organism	Structure
additional information	-	additional information	steady-state kinetics	Methylosinus sporium

Metals/Ions

Metals/Ions	Comment	Organism	Structure
Cu2+	metal-binding titrations of MMOD and copper	Methylosinus sporium
Fe	structural comparison of the di-iron center of MMOH (PDB ID 1MHY), MMOH-MMOB (PDB ID 4GAM), and MMOH-MMOD. Conformational changes in the MMOH four-helix bundle (helices B, C, E, and F) upon MMOB and MMOD binding. Six residues that coordinate to the two iron atoms are distributed on to the different helices	Methylosinus sporium

Natural Substrates/ Products (Substrates)

Natural Substrates	Organism	Comment (Nat. Sub.)	Natural Products	Comment (Nat. Pro.)	Rev.
methane + NADH + H+ + O2	Methylosinus sporium	-	methanol + NAD+ + H2O	-	?
methane + NADH + H+ + O2	Methylosinus sporium 5	-	methanol + NAD+ + H2O	-	?
methane + NADH + H+ + O2	Methylosinus sporium ATCC 35069	-	methanol + NAD+ + H2O	-	?

Organism

Organism	UniProt	Comment	Textmining
Methylosinus sporium	-	-	-
Methylosinus sporium 5	-	-	-
Methylosinus sporium ATCC 35069	-	-	-

Purification (Commentary)

Purification (Comment)	Organism
native enzyme complex from cell culture by two different steps of anion exchange chromatography, recombinant subunits MMOB and MMOR from Escherichia coli strain BL21(DE3) by two different steps of anion exchange chromatography, and gel filtration, recombinant His-MBP-tagged MMOD from Escherichia coli strain Rosetta (DE3) by nickel and amylose affinity chromatography, the tag is cleaved off by tobacco etch virus protease followed by anion exchange chromatography and gel filtration	Methylosinus sporium

Purification (Comment)

Organism

native enzyme complex from cell culture by two different steps of anion exchange chromatography, recombinant subunits MMOB and MMOR from Escherichia coli strain BL21(DE3) by two different steps of anion exchange chromatography, and gel filtration, recombinant His-MBP-tagged MMOD from Escherichia coli strain Rosetta (DE3) by nickel and amylose affinity chromatography, the tag is cleaved off by tobacco etch virus protease followed by anion exchange chromatography and gel filtration

Methylosinus sporium

Substrates and Products (Substrate)

Substrates	Comment Substrates	Organism	Products	Comment (Products)	Rev.
methane + NADH + H+ + O2	-	Methylosinus sporium	methanol + NAD+ + H2O	-	?
methane + NADH + H+ + O2	-	Methylosinus sporium 5	methanol + NAD+ + H2O	-	?
methane + NADH + H+ + O2	-	Methylosinus sporium ATCC 35069	methanol + NAD+ + H2O	-	?
additional information	sMMO is known to oxidize a variety of hydrocarbons, including alkanes ranging from methane to octane. The presence of 1,6-hexanediol near the di-iron center can be explained by the opening of the cavity, mediated by the side-chain rearrangement of Leu110 and Phe188, both of which function together as a gate for substrate and product passage to the active site. While MMOB is known to connect cavities for substrate access, the MMOD-mediated cavity opening appears to be a consequence of MMOHbeta-NT dissociation and subsequent structural relaxation of MMOHalpha. Both substrate ingress and product egress may take place through the substrate access cavity and not through the pore located near the active site, at least for hydrocarbon chain substrates such as hexane	Methylosinus sporium	?	-	-
additional information	sMMO is known to oxidize a variety of hydrocarbons, including alkanes ranging from methane to octane. The presence of 1,6-hexanediol near the di-iron center can be explained by the opening of the cavity, mediated by the side-chain rearrangement of Leu110 and Phe188, both of which function together as a gate for substrate and product passage to the active site. While MMOB is known to connect cavities for substrate access, the MMOD-mediated cavity opening appears to be a consequence of MMOHbeta-NT dissociation and subsequent structural relaxation of MMOHalpha. Both substrate ingress and product egress may take place through the substrate access cavity and not through the pore located near the active site, at least for hydrocarbon chain substrates such as hexane	Methylosinus sporium 5	?	-	-
additional information	sMMO is known to oxidize a variety of hydrocarbons, including alkanes ranging from methane to octane. The presence of 1,6-hexanediol near the di-iron center can be explained by the opening of the cavity, mediated by the side-chain rearrangement of Leu110 and Phe188, both of which function together as a gate for substrate and product passage to the active site. While MMOB is known to connect cavities for substrate access, the MMOD-mediated cavity opening appears to be a consequence of MMOHbeta-NT dissociation and subsequent structural relaxation of MMOHalpha. Both substrate ingress and product egress may take place through the substrate access cavity and not through the pore located near the active site, at least for hydrocarbon chain substrates such as hexane	Methylosinus sporium ATCC 35069	?	-	-

Synonyms

Synonyms	Comment	Organism
sMMO	-	Methylosinus sporium
soluble methane monooxygenase	-	Methylosinus sporium

Temperature Optimum [°C]

Temperature Optimum [°C]	Temperature Optimum Maximum [°C]	Comment	Organism
25	-	assay at	Methylosinus sporium

pH Optimum

pH Optimum Minimum	pH Optimum Maximum	Comment	Organism
7.5	-	assay at	Methylosinus sporium

Cofactor

Cofactor	Comment	Organism	Structure
FAD	-	Methylosinus sporium
NADH	-	Methylosinus sporium

General Information

General Information	Comment	Organism
additional information	structure comparisons of the enzymes from Methylosinus sporium strain 5 and Methylosinus trichosporium strain OB3b. MMOH-MMOD complex modeling, overview	Methylosinus sporium
physiological function	soluble methane monooxygenase in methanotrophs converts methane to methanol under ambient conditions. The maximum catalytic activity of hydroxylase (MMOH) is achieved through the interplay of its regulatory protein (MMOB) and reductase. An additional auxiliary protein, MMOD, functions as an inhibitor for MMOH by competing with MMOB for MMOH association as well as by disrupting the active geometric form of the di-iron center. The expression level of MMOD is relatively low and it binds tightly with MMOH near the di-iron center. ApoMMOH (iron removed) in the presence of MMOD or MMOB demonstrates that both MMOD and MMOB block iron loading toward apoMMOH instead of promoting it. Both iron atoms show full occupancy at the di-iron center during structure refinement, indicating that there is no loss of iron upon MMOD association. One potential function is that MMOD acts as a protein chaperone to assist the protein folding of MMOH by protecting MMOH until MMOHbeta-NT latches on as the final step of the protein folding process, potential function of MMOD as a protein chaperone	Methylosinus sporium