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Inactivation of the particulate methane monooxygenase (pMMO) in Methylococcus capsulatus (Bath) by acetylene

Pham, Minh D. and Lin, Ya-Ping and Vuong, Quan Van and Nagababu, Penumaka and Chang, Brian T.-A. and Ng, Kok Yaoh and Chen, Chein-Hung and Han, Chau-Chung and Chen, Chung-Hsuan and Li, Mai Suan and Yu, Steve S.-F. and Chan, Sunney I. (2015) Inactivation of the particulate methane monooxygenase (pMMO) in Methylococcus capsulatus (Bath) by acetylene. Biochimica et Biophysica Acta - Proteins and Proteomics, 1854 (12). pp. 1842-1852. ISSN 1570-9639.

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Acetylene (HCCH) has a long history as a mechanism-based enzyme inhibitor and is considered an active-site probe of the particulate methane monooxygenase (pMMO). Here, we report how HCCH inactivates pMMO in Methylococcus capsulatus (Bath) by using high-resolution mass spectrometry and computational simulation. High-resolution MALDI-TOF MS of intact pMMO complexes has allowed us to confirm that the enzyme oxidizes HCCH to the ketene (C_2H_2O) intermediate, which then forms an acetylation adduct with the transmembrane PmoC subunit. LC-MS/MS analysis of the peptides derived from in-gel proteolytic digestion of the protein subunit identifies K196 of PmoC as the site of acetylation. No evidence is obtained for chemical modification of the PmoA or PmoB subunit. The inactivation of pMMO by a single adduct in the transmembrane PmoC domain is intriguing given the complexity of the structural fold of this large membrane-protein complex as well as the complicated roles played by the various metal cofactors in the enzyme catalysis. Computational studies suggest that the entry of hydrophobic substrates to, and migration of products from, the catalytic site of pMMO is controlled tightly within the transmembrane domain. Support of these conclusions is provided by parallel experiments with two related alkynes: propyne (CH3CCH) and trifluoropropyne (CF_3CCH). Finally, we discuss the implication of these findings to the location of the catalytic site in pMMO.

Item Type:Article
Related URLs:
URLURL TypeDescription
Yu, Steve S.-F.0000-0002-3462-065X
Chan, Sunney I.0000-0002-5348-2723
Additional Information:© 2015 Elsevier B.V. Received date: 16 June 2015; Revised date: 29 July 2015; Accepted date: 9 August 2015. This work was supported by Academia Sinica and by research grants from the National Science Council of the Republic of China (NSC 98-2113-M-001-026; NSC 99-2119-M-001-003; NSC 100-2113-M-001-002; and 101-2113-M-001-013- to SIC; and NSC 101-2113-M-001-007-MY3 to SSFY). The computer simulations were supported by the Department of Science and Technology at Ho Chi Minh City, Vietnam and Narodowe Centrum Nauki in Poland (grant 2011/01/B/NZ1/01622 to MSL). We acknowledge numerous scientific discussions on this work with Dr. Chia-Min Yang (Department of Chemistry, National Tsing Hua University, Hsinchu, Taiwan). M.D.P. is also grateful to Dr. Yang for serving as his academic advisor.
Funding AgencyGrant Number
Academia SinicaUNSPECIFIED
National Science Council (Taipei)NSC 98-2113-M-001-026
National Science Council (Taipei)NSC 99-2119-M-001-003
National Science Council (Taipei)NSC 100-2113-M-001-002
National Science Council (Taipei)NSC 101-2113-M-001-013
National Science Council (Taipei)NSC 101-2113-M-001-007-MY3
Department of Science and Technology (Vietnam)UNSPECIFIED
National Science Centre (Poland)2011/01/B/NZ1/01622
Subject Keywords:Acetylene; Mechanism-based inactivation; Particulate methane monooxygenase; Methylococcus capsulatus (Bath); Mass spectrometry; Computational simulation
Issue or Number:12
Record Number:CaltechAUTHORS:20150824-122929840
Persistent URL:
Official Citation:Minh D. Pham, Ya-Ping Lin, Quan Van Vuong, Penumaka Nagababu, Brian T.-A. Chang, Kok Yaoh Ng, Chein-Hung Chen, Chau-Chung Han, Chung-Hsuan Chen, Mai Suan Li, Steve S.-F. Yu, Sunney I. Chan, Inactivation of the particulate methane monooxygenase (pMMO) in Methylococcus capsulatus (Bath) by acetylene, Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics, Volume 1854, Issue 12, December 2015, Pages 1842-1852, ISSN 1570-9639, (
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:59845
Deposited By: Tony Diaz
Deposited On:24 Aug 2015 22:59
Last Modified:09 Mar 2020 13:19

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