PMID- 17154527
OWN - NLM
STAT- MEDLINE
DCOM- 20070221
LR  - 20131121
IS  - 0006-2960 (Print)
IS  - 0006-2960 (Linking)
VI  - 45
IP  - 50
DP  - 2006 Dec 19
TI  - Altering conserved lipid binding sites in cytochrome c oxidase of Rhodobacter 
      sphaeroides perturbs the interaction between subunits I and III and promotes 
      suicide inactivation of the enzyme.
PG  - 14896-907
AB  - Subunit III of the three-subunit catalytic core of cytochrome c oxidase (CcO) 
      contains no metal centers, but it does bind two lipids, within a deep cleft, in 
      binding sites conserved from bacteria to humans. Subunit III binds to subunit I, 
      where it prevents the spontaneous suicide inactivation of CcO by decreasing the 
      probability of side reactions at the heme-Cu O2 reduction site in subunit I. 
      Subunit III prevents suicide inactivation by (1) maintaining adequate rates of 
      proton delivery to the heme-Cu active site and (2) stabilizing the structure of 
      the active site during turnover [Mills and Hosler (2005) Biochemistry 44, 4656]. 
      Here, we first show that mutating several individual residues of the conserved 
      lipid binding sites in subunit III disturbs the subunit I-III interface. Then, 
      two lipid binding site mutants were constructed with an affinity tag on subunit 
      III such that the mutant CcOs could be isolated with 100% subunit III. R226A 
      eliminates an ion pair to the phosphate of the outermost lipid of the cleft, 
      while W59A-F86A disrupts interactions with the fatty acid tails of both lipids. 
      Once these mutant CcOs are placed into soybean phospholipid vesicles, where 
      extensive exchange of bacterial for soybean lipids takes place, it is shown that 
      altering the lipid binding sites mimics a major loss of subunit III, even though 
      subunit III is completely retained, in that suicide inactivation becomes much 
      more probable. The rate of proton delivery to the active site remains rapid, 
      ruling out slow proton uptake as the primary reason for increased suicide 
      inactivation upon alteration of the lipid binding sites. We conclude that 
      altering the lipid binding sites of subunit III may promote side reactions 
      leading to suicide inactivation by allowing greater movement to occur in and 
      around the O2 reduction site of subunit I during the catalytic cycle.
FAU - Varanasi, Lakshman
AU  - Varanasi L
AD  - Department of Biochemistry, University of Mississippi Medical Center, 2500 North 
      State Street, Jackson, Mississippi 39216, USA.
FAU - Mills, Denise
AU  - Mills D
FAU - Murphree, Anna
AU  - Murphree A
FAU - Gray, Jimmy
AU  - Gray J
FAU - Purser, Chris
AU  - Purser C
FAU - Baker, Rodney
AU  - Baker R
FAU - Hosler, Jonathan
AU  - Hosler J
LA  - eng
GR  - GM26916/GM/NIGMS NIH HHS/United States
GR  - GM56824/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, N.I.H., Extramural
PL  - United States
TA  - Biochemistry
JT  - Biochemistry
JID - 0370623
RN  - 0 (Bacterial Proteins)
RN  - 0 (Lipids)
RN  - 0 (Protein Subunits)
RN  - 789U1901C5 (Copper)
RN  - EC 1.9.3.1 (Electron Transport Complex IV)
RN  - S88TT14065 (Oxygen)
SB  - IM
MH  - Amino Acid Substitution
MH  - Bacterial Proteins/*chemistry/genetics/metabolism
MH  - Binding Sites/genetics
MH  - Catalysis
MH  - Copper/chemistry/metabolism
MH  - Electron Transport Complex IV/*chemistry/genetics/metabolism
MH  - *Enzyme Activation
MH  - Lipids/*chemistry
MH  - Oxidation-Reduction
MH  - Oxygen/chemistry/metabolism
MH  - Protein Binding/genetics
MH  - Protein Structure, Quaternary
MH  - Protein Subunits/genetics
MH  - Rhodobacter sphaeroides/*enzymology/genetics
EDAT- 2006/12/13 09:00
MHDA- 2007/02/22 09:00
CRDT- 2006/12/13 09:00
PHST- 2006/12/13 09:00 [pubmed]
PHST- 2007/02/22 09:00 [medline]
PHST- 2006/12/13 09:00 [entrez]
AID - 10.1021/bi061390q [doi]
PST - ppublish
SO  - Biochemistry. 2006 Dec 19;45(50):14896-907. doi: 10.1021/bi061390q.