RESUMO
An a priori model-independent method for the determination of accurate spectra of photocycle intermediates is developed. The method, singular value decomposition with self-modeling (SVD-SM), is tested on simulated difference spectra designed to mimic the photocycle of the Asp-96 --> Asn mutant of bacteriorhodopsin. Stoichiometric constraints, valid until the onset of the recovery of bleached bacteriorhodopsin at the end of the photocycle, guide the self-modeling procedure. The difference spectra of the intermediates are determined in eigenvector space by confining the search for their coordinates to a stoichiometric plane. In the absence of random noise, SVD-SM recovers the intermediate spectra and their time evolution nearly exactly. The recovery of input spectra and kinetics is excellent although somewhat less exact when realistic random noise is included in the input spectra. The difference between recovered and input kinetics is now visually discernible, but the same reaction scheme with nearly identical rate constants to those assumed in the simulation fits the output kinetics well. SVD-SM relegates the selection of a photocycle model to the late stage of the analysis. It thus avoids derivation of erroneous model-specific spectra that result from global model-fitting approaches that assume a model at the outset.
Assuntos
Bacteriorodopsinas/química , Bacteriorodopsinas/metabolismo , Bacteriorodopsinas/efeitos da radiação , Cinética , Modelos Teóricos , Fotoquímica , Espectrofotometria , Fatores de TempoRESUMO
Singular value decomposition with self-modeling is applied to resolve the intermediate spectra and kinetics of the Asp96 --> Asn mutant bacteriorhodopsin. The search for the difference spectra of the intermediates is performed in eigenvector space on the stoichiometric plane. The analysis of data at pH values ranging from 4 to 8 and temperatures between 5 and 25 degrees C reveals significant, early partial recovery of the initial state after photoexcitation. The derived spectra are not biased by assumed photocycles. The intermediate spectra derived in the initial step differ from spectra determined in prior analyses, which results in intermediate concentrations with improved stoichiometric properties. Increasingly more accurate photocycles follow with increasing assumed complexity, of which parallel models are favored, consistent with recent, independent experimental evidence.
Assuntos
Asparagina , Ácido Aspártico , Bacteriorodopsinas/química , Bacteriorodopsinas/metabolismo , Substituição de Aminoácidos , Halobacterium salinarum/metabolismo , Cinética , Modelos Químicos , Mutagênese Sítio-Dirigida , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismoAssuntos
Pulmão/efeitos dos fármacos , Adulto , Dióxido de Carbono/metabolismo , Criança , Feminino , Humanos , Pulmão/fisiologia , Masculino , Métodos , Consumo de Oxigênio/efeitos dos fármacos , Pressão , Alvéolos Pulmonares/efeitos dos fármacos , Respiração/efeitos dos fármacos , Fumar , Espirometria , Temperatura , Fatores de TempoAssuntos
Ácidos Graxos , Ureia , Coloides , Eletroquímica , Ácidos Palmíticos , Ácidos Esteáricos , Propriedades de Superfície , TemperaturaAssuntos
Gases/farmacologia , Hidrocarbonetos , Óleos , Pressão , Anestesia , Argônio/farmacologia , Dióxido de Carbono/farmacologia , Membrana Celular/fisiologia , Eletroquímica , Hélio/farmacologia , Nitrogênio/farmacologia , Óxido Nitroso/farmacologia , Pressão Parcial , Propriedades de Superfície , Xenônio/farmacologiaRESUMO
Diffusion of the chloride salts of Li, Na, K, Rb, and Cs from water into 1-butanol and from 1-butanol into water was examined at temperatures from 13-40 degrees C. Distribution coefficients, interfacial transfer coefficients, Arrhenius activation energy, free energy of activation, enthalpy, and entropy of activation were determined for the diffusion of these salts across the alcohol-water interface. The results indicate that the entropy decrease made the major contribution to the change in the free energy of activation.