PMID- 19243111
OWN - NLM
STAT- MEDLINE
DCOM- 20100405
LR  - 20131121
IS  - 1520-4995 (Electronic)
IS  - 0006-2960 (Linking)
VI  - 48
IP  - 11
DP  - 2009 Mar 24
TI  - Microscopic pKa analysis of Glu286 in cytochrome c oxidase (Rhodobacter 
      sphaeroides): toward a calibrated molecular model.
PG  - 2468-85
LID - 10.1021/bi8021284 [doi]
AB  - As stringent tests for the molecular model and computational protocol, 
      microscopic pK(a) calculations are performed for the key residue, Glu286, in 
      cytochrome c oxidase (CcO) using a combined quantum mechanical/molecular 
      mechanical (QM/MM) potential and a thermodynamic integration protocol. The impact 
      of the number of water molecules in the hydrophobic cavity and protonation state 
      of several key residues (e.g., His334, Cu(B)-bound water, and PRD(a3)) on the 
      computed microscopic pK(a) values of Glu286 has been systematically examined. To 
      help evaluate the systematic errors in the QM/MM-based protocol, microscopic 
      pK(a) calculations have also been carried out for sites in a soluble protein 
      (Asp70 in T4 lysozyme) and a better-characterized membrane protein (Asp85 in 
      bacteriorhodopsin). Overall, the results show a significant degree of internal 
      consistency and reproducibility that support the effectiveness of the 
      computational framework. Although the number of water molecules in the 
      hydrophobic cavity does not greatly influence the computed pK(a) of Glu286, the 
      protonation states of several residues, some of which are rather far away, have 
      more significant impacts. Adopting the standard protonation state for all 
      titratable residues leaves a large net charge on the system and a significantly 
      elevated pK(a) for Glu286, highlighting that any attempt to address the 
      energetics of proton transfers in CcO at a microscopic level should carefully 
      select the protonation state of residues, even those not in the immediate 
      neighborhood of the active site. The calculations indirectly argue against the 
      deprotonation of His334 for the proton pumping process, although further studies 
      that explicitly compute its pK(a) are required for a more conclusive statement. 
      Finally, the deprotonated Glu286 is found to be in a stable water-mediated 
      connection with PRD(a3) for at least several nanoseconds when this presumed 
      pumping site is protonated. This does not support the proposed role of Glu286 as 
      a robust gating valve that prevents proton leakage, although a conclusive 
      statement awaits a more elaborate characterization of the Glu286-PRD(a3) 
      connectivity with free energy simulations and a protonated PRD(a3). The large 
      sets of microscopic simulations performed here have provided useful guidance to 
      the establishment of a meaningful molecular model and effective computational 
      protocol for explicitly analyzing the proton transfer kinetics in CcO, which is 
      required for answering key questions regarding the pumping function of this 
      fascinating and complex system.
FAU - Ghosh, Nilanjan
AU  - Ghosh N
AD  - Department of Chemistry, University of Wisconsin, 1101 University Avenue, 
      Madison, Wisconsin 53706, USA.
FAU - Prat-Resina, Xavier
AU  - Prat-Resina X
FAU - Gunner, M R
AU  - Gunner MR
FAU - Cui, Qiang
AU  - Cui Q
LA  - eng
SI  - PDB/1M56
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PT  - Research Support, U.S. Gov't, Non-P.H.S.
PL  - United States
TA  - Biochemistry
JT  - Biochemistry
JID - 0370623
RN  - 0 (Bacterial Proteins)
RN  - 0RH81L854J (Glutamine)
RN  - EC 1.9.3.1 (Electron Transport Complex IV)
SB  - IM
MH  - Bacterial Proteins/*chemistry
MH  - Computer Simulation
MH  - Electron Transport Complex IV/*chemistry
MH  - Glutamine/chemistry
MH  - Kinetics
MH  - Models, Molecular
MH  - Molecular Conformation
MH  - Rhodobacter sphaeroides/chemistry/*enzymology
MH  - Thermodynamics
EDAT- 2009/02/27 09:00
MHDA- 2010/04/07 06:00
CRDT- 2009/02/27 09:00
PHST- 2009/02/27 09:00 [entrez]
PHST- 2009/02/27 09:00 [pubmed]
PHST- 2010/04/07 06:00 [medline]
AID - 10.1021/bi8021284 [doi]
PST - ppublish
SO  - Biochemistry. 2009 Mar 24;48(11):2468-85. doi: 10.1021/bi8021284.