Atopy, asthma, and lung function in relation to folate and vitamin B(12) in adults.

Background: Recent studies suggested low serum folate and impaired folate metabolism as potential risk factors for development of asthma and atopic disease, but the results are inconsistent. The aim of this study was to investigate the relations of markers of folate and vitamin B₁₂ (B₁₂) deficiency with different phenotypes of asthma and atopy.

Methods: A random sample of 6784 persons from a general population aged 30-60 years participated in a health examination in 1999-2001, and 4516 (66.

A mechanistic principle for proton pumping by cytochrome c oxidase.

In aerobic organisms, cellular respiration involves electron transfer to oxygen through a series of membrane-bound protein complexes. The process maintains a transmembrane electrochemical proton gradient that is used, for example, in the synthesis of ATP. In mitochondria and many bacteria, the last enzyme complex in the electron transfer chain is cytochrome c oxidase (CytcO), which catalyses the four-electron reduction of O2 to H2O using electrons delivered by a water-soluble donor, cytochrome c.

Subunit III of cytochrome c oxidase of Rhodobacter sphaeroides is required to maintain rapid proton uptake through the D pathway at physiologic pH.

The catalytic core of cytochrome c oxidase is composed of three subunits where subunits I and II contain all of the redox-active metal centers and subunit III is a seven transmembrane helix protein that binds to subunit I. The N-terminal region of subunit III is adjacent to D132 of subunit I, the initial proton acceptor of the D pathway that transfers protons from the protein surface to the buried active site approximately 30 A distant. The absence of subunit III only slightly alters the initial steady-state activity of the oxidase at pH 6.

Relocation of an internal proton donor in cytochrome c oxidase results in an altered pK(a) and a non-integer pumping stoichiometry.

Cytochrome c oxidase from Rhodobacter sphaeroides has two proton-input pathways leading from the protein surface towards the catalytic site, located within the membrane-spanning part of the enzyme. One of these pathways, the D-pathway, contains a highly conserved Glu residue [E(I-286)], which plays an important role in proton transfer through the pathway. In a recent study, we showed that a mutant enzyme in which E(I-286) was re-located to the opposite side of the D-pathway [EA(I-286)/IE(I-112) double mutant enzyme] was able to pump protons, although with a stoichiometry that was lower than that of the wild-type enzyme (approximately 0.

Redesign of the proton-pumping machinery of cytochrome c oxidase: proton pumping does not require Glu(I-286).

One of the putative proton-transfer pathways leading from solution toward the binuclear center in many cytochrome c oxidases is the D-pathway, so-called because it starts with a highly conserved aspartate [D(I-132)] residue. Another highly conserved amino acid residue in this pathway, glutamate(I-286), has been indicated to play a central role in the proton-pumping machinery of mitochondrial-type enzymes, a role that requires a movement of the side chain between two distinct positions. In the present work we have relocated the glutamate to the opposite side of the proton-transfer pathway by constructing the double mutant EA(I-286)/IE(I-112).

Kinetics of electron and proton transfer during O(2) reduction in cytochrome aa(3) from A. ambivalens: an enzyme lacking Glu(I-286).

Acidianus ambivalens is a hyperthermoacidophilic archaeon which grows optimally at approximately 80 degrees C and pH 2.5. The terminal oxidase of its respiratory system is a membrane-bound quinol oxidase (cytochrome aa(3)) which belongs to the heme-copper oxidase superfamily.

Proton transfer from glutamate 286 determines the transition rates between oxygen intermediates in cytochrome c oxidase.

We have investigated the electron-proton coupling during the peroxy (P(R)) to oxo-ferryl (F) and F to oxidised (O) transitions in cytochrome c oxidase from Rhodobacter sphaeroides. The kinetics of these reactions were investigated in two different mutant enzymes: (1) ED(I-286), in which one of the key residues in the D-pathway, E(I-286), was replaced by an aspartate which has a shorter side chain than that of the glutamate and, (2) ML(II-263), in which the redox potential of Cu(A) is increased by approximately 100 mV, which slows electron transfer to the binuclear centre during the F-->O transition by a factor of approximately 200. In ED(I-286) proton uptake during P(R)-->F was slowed by a factor of approximately 5, which indicates that E(I-286) is the proton donor to P(R).

Dynamics of the binuclear center of the quinol oxidase from Acidianus ambivalens.

We have investigated the kinetic and thermodynamic properties of carbon monoxide binding to the fully reduced quinol oxidase (cytochrome aa(3)) from the hyperthermophilic archaeon Acidianus ambivalens. After flash photolysis of CO from heme a(3), the complex recombines with an apparent rate constant of approximately 3 s(-1), which is much slower than with the bovine cytochrome c oxidase (approximately 80 s(-1)). Investigation of the CO-recombination rate as a function of the CO concentration shows that the rate saturates at high CO concentrations, which indicates that CO must bind transiently to Cu(B) before binding to heme a(3).