Structural characterization by EXAFS spectroscopy of the binuclear iron center in protein A of methane monooxygenase from Methylococcus capsulatus (Bath)

1988 ◽  
Vol 110 (7) ◽  
pp. 2330-2332 ◽  
Author(s):  
Agneta. Ericson ◽  
Britt. Hedman ◽  
Keith O. Hodgson ◽  
Jeffrey. Green ◽  
Howard. Dalton ◽  
...  
Keyword(s):  
Structural Characterization ◽  
Methane Monooxygenase ◽  
Protein A ◽  
Exafs Spectroscopy ◽  
Methylococcus Capsulatus ◽  
Binuclear Iron ◽  
Iron Center

Optimization of solubilization and purification procedures for the hydroxylase component of membrane-bound methane monooxygenase from Methylococcus capsulatus strain M

2006 ◽  
Vol 71 (12) ◽  
pp. 1329-1335 ◽  
Author(s):  
V. I. Vasil’ev ◽  
T. V. Tikhonova ◽  
R. I. Gvozdev ◽  
I. A. Tukhvatullin ◽  
V. O. Popov
Keyword(s):  
Methane Monooxygenase ◽  
Methylococcus Capsulatus ◽  
Membrane Bound

Structural Characterization of Organocuprate reagents. EXAFS Spectroscopy and ab Initio Calculations

1995 ◽  
Vol 117 (50) ◽  
pp. 12489-12497 ◽  
Author(s):  
Timothy L. Stemmler ◽  
Terence M. Barnhart ◽  
James E. Penner-Hahn ◽  
Charles E. Tucker ◽  
Paul Knochel ◽  
...  
Keyword(s):  
Ab Initio Calculations ◽  
Ab Initio ◽  
Structural Characterization ◽  
Exafs Spectroscopy ◽  
Organocuprate Reagents

scholarly journals A stopped-flow kinetic study of soluble methane mono-oxygenase from Methylococcus capsulatus (Bath)

1989 ◽  
Vol 259 (1) ◽  
pp. 167-172 ◽  
Author(s):  
J Green ◽  
H Dalton
Keyword(s):  
Electron Transfer ◽  
Protein C ◽  
Turnover Rate ◽  
Protein A ◽  
Methylococcus Capsulatus ◽  
Bond Breakage ◽  
Conversion Of Methane ◽  
Protein Components ◽  
A Charge ◽  
Rate Limiting

1. The roles of the three protein components of soluble methane mono-oxygenase were investigated by the use of rapid-reaction techniques. The transfer of electrons through the enzyme complex from NADH to methane/O2 was also investigated. 2. Electron transfer from protein C, the reductase component, to protein A, the hydroxylase component, was demonstrated. Protein C was shown to undergo a three-electron--one-electron catalytic cycle. The interaction of protein C with NADH was investigated. Reduction of protein C was shown to be rapid, and a charge-transfer interaction between reduced FAD and NAD+ was observed; this intermediate was also found in static titration experiments. Thus the binding of NADH, the reduction of protein C and the intramolecular transfer of electrons through protein C were shown to be much more rapid than the turnover rate of methane mono-oxygenase. 3. The rate of transfer of electrons from protein C to protein A was shown to be lower than the reduction of protein C but higher than the turnover rate of methane mono-oxygenase. Association of the proteins was not rate-limiting. The amount of protein A present in the system had a small effect on the rate of reduction of protein C, indicating some co-operativity between the two proteins. 4. Protein B was shown to prevent electron transfer between protein C and protein A in the absence of methane. On addition of saturating concentrations of methane electron transfer was restored. With saturating concentrations of methane and O2 the observed rate constant for the conversion of methane into methanol was 0.26 s-1 at 18 degrees C. 5. By the use of [2H4]methane it was demonstrated that C-H-bond breakage is likely to be the rate-limiting step in the conversion of methane into methanol.

scholarly journals Expression of Individual Copies ofMethylococcus capsulatus Bath Particulate Methane Monooxygenase Genes

2001 ◽  
Vol 183 (5) ◽  
pp. 1810-1812 ◽  
Author(s):  
Sergei Stolyar ◽  
Marion Franke ◽  
Mary E. Lidstrom
Keyword(s):  
Methane Monooxygenase ◽  
Gene Clusters ◽  
Gene Fusions ◽  
Methylococcus Capsulatus ◽  
Chromosomal Gene ◽  
Particulate Methane Monooxygenase ◽  
Relative Expression

ABSTRACT The expression of the two gene clusters encoding the particulate methane monooxygenase (pMMO) in Methylococcus capsulatusBath was assessed by analysis of transcripts and by use of chromosomal gene fusions. The results suggest that the two clusters are functionally redundant but that relative expression alters depending on the copper levels available for growth.

scholarly journals Batch Production of High-Quality Graphene Grids for Cryo-EM: Cryo-EM Structure of Methylococcus capsulatus Soluble Methane Monooxygenase Hydroxylase

2021 ◽  
Author(s):  
Eungjin Ahn ◽  
byungchul Kim ◽  
uhn-soo Cho
Keyword(s):  
Protein Denaturation ◽  
Methane Monooxygenase ◽  
Atomic Layer ◽  
Water Interface ◽  
Methylococcus Capsulatus ◽  
Force Microscopy ◽  
Voltage Electron ◽  
Soluble Methane Monooxygenase ◽  
Air Water Interface ◽  
Scanning Em

Cryogenic electron microscopy (cryo-EM) has become a widely used tool for determining protein structure. Despite recent advances in instruments and algorithms, sample preparation remains a major bottleneck for several reasons, including protein denaturation at the air/water interface and the presence of preferred orientations and nonuniform ice layers. Graphene, a two-dimensional allotrope of carbon consisting of a single atomic layer, has recently attracted attention as a near-ideal support film for cryo-EM that can overcome these challenges because of its superior properties, including mechanical strength and electrical conductivity. Graphene minimizes background noise and provides a stable platform for specimens under a high-voltage electron beam and cryogenic conditions. Here, we introduce a reliable, easily implemented, and reproducible method of producing 36 graphene-coated grids at once within 1.5 days. The quality of the graphene grids was assessed using various tools such as scanning EM, Raman spectroscopy, and atomic force microscopy. To demonstrate their practical application, we determined the cryo-EM structure of Methylococcus capsulatus soluble methane monooxygenase hydroxylase (sMMOH) at resolutions of 2.9 and 2.4 angstrom using Quantifoil and graphene-coated grids, respectively. We found that the graphene-coated grid has several advantages; for example, it requires less protein, enables easy control of the ice thickness, and prevents pro-tein denaturation at the air/water interface. By comparing the cryo-EM structure of sMMOH with its crystal structure, we revealed subtle yet significant geometrical differences at the non-heme di-iron center, which may better indicate the active site configuration of sMMOH in the resting/oxidized state.

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