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A thermodynamic analysis of the interaction between the mitochondrial coupling adenosine triphosphatase and its naturally occurring inhibitor protein.

ABSTRACT: 1. The naturally occurring ATPase (adenosine triphosphatase)-inhibitor protein, from bovine heart mitochondria, was obtained as a single pure protein. It was not identical with any of the five subunits (alpha-epsilon) of the isolated ATPase, and appeared to be a single polypeptide chain. 2. The inhibitor combined with the ATPase in a 1:1 molar ratio, producing a completely inhibited ATPase molecule. The affinity of the ATPase for its inhibitor is high; the K(d) is of the order of 10(-8)m. 3. The enthalpy of the ATPase-inhibitor complex-formation is positive, the value of K(d) decreasing as the temperature is raised. This suggests that the forces involved are largely hydrophobic in nature. 4. Hydrolysis of a nucleoside triphosphate promoted formation of the ATPase-inhibitor complex, although the equilibrium position was almost unaffected by the rate of hydrolysis. At low salt concentration, less than 200 turnovers of the ATPase suffice for the ATPase to combine with the inhibitor protein. At higher salt concentrations, a larger number of turnovers is required. It is suggested that the inhibitor binds to a form of the ATPase that is produced transiently during hydrolysis. 5. In the presence of 75mm-K(2)SO(4), the rates of association and dissociation are slow enough to allow their kinetics to be studied. Association is first-order in inhibitor concentration, but fractional order in ATPase concentration. Dissociation is first-order in ATPase-inhibitor complex concentration. The temperature coefficients of the ;on' and ;off' processes were also measured. 6. A simple kinetic model for the ATPase-inhibitor interaction is proposed that can be extended to take into account release of inhibitor protein under energized conditions on the membrane. 7. The isolated ATPase is inhibited by preincubation with Mg(2+), reversible by subsequent addition of EDTA, and by ADP, reversible by subsequent addition of ATP. These effects are not found on the membrane-bound ATPase. The mechanism of these effects is discussed.

SUBMITTER: Gomez-Fernandez JC

PROVIDER: S-EPMC1186322 | biostudies-other | 1978 Dec

REPOSITORIES: biostudies-other

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Project description:Here, we examine in silico the infection dynamics and interactions of two Zika virus (ZIKV) genomes: one is the full-length ZIKV genome (wild type [WT]), and the other is one of the naturally occurring defective viral genomes (DVGs), which can replicate in the presence of the WT genome, appears under high-MOI (multiplicity of infection) passaging conditions, and carries a deletion encompassing part of the structural and NS1 protein-coding region. Ordinary differential equations (ODEs) were used to simulate the infection of cells by virus particles and the intracellular replication of the WT and DVG genomes that produce these particles. For each virus passage in Vero and C6/36 cell cultures, the rates of the simulated processes were fitted to two types of observations: virus titer data and the assembled haplotypes of the replicate passage samples. We studied the consistency of the model with the experimental data across all passages of infection in each cell type separately as well as the sensitivity of the model's parameters. We also determined which simulated processes of virus evolution are the most important for the adaptation of the WT and DVG interplay in these two disparate cell culture environments. Our results demonstrate that in the majority of passages, the rates of DVG production are higher inC6/36 cells than in Vero cells, which might result in tolerance and therefore drive the persistence of the mosquito vector in the context of ZIKV infection. Additionally, the model simulations showed a slower accumulation of infected cells under higher activation of the DVG-associated processes, which indicates a potential role of DVGs in virus attenuation. IMPORTANCE One of the ideas for lessening Zika pathogenicity is the addition of its natural or engineered defective virus genomes (DVGs) (have no pathogenicity) to the infection pool: a DVG is redirecting the wild-type (WT)-associated virus development resources toward its own maturation. The mathematical model presented here, attuned to the data from interplays between WT Zika viruses and their natural DVGs in mammalian and mosquito cells, provides evidence that the loss of uninfected cells is attenuated by the DVG development processes. This model enabled us to estimate the rates of virus development processes in the WT/DVG interplay, determine the key processes, and show that the key processes are faster in mosquito cells than in mammalian ones. In general, the presented model and its detailed study suggest in what important virus development processes the therapeutically efficient DVG might compete with the WT; this may help in assembling engineered DVGs for ZIKV and other flaviviruses.

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A thermodynamic analysis of the interaction between the mitochondrial coupling adenosine triphosphatase and its naturally occurring inhibitor protein.

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