Assuntos
Anti-Infecciosos Urinários/urina , Piridazinas/urina , Administração Oral , Alcalose/metabolismo , Cloreto de Amônio/farmacologia , Animais , Anti-Infecciosos Urinários/administração & dosagem , Anti-Infecciosos Urinários/sangue , Bicarbonatos/farmacologia , Disponibilidade Biológica , Dioxolanos/administração & dosagem , Dioxolanos/sangue , Dioxolanos/urina , Cães , Feminino , Humanos , Concentração de Íons de Hidrogênio , Técnicas In Vitro , Cinética , Piridazinas/administração & dosagem , Piridazinas/sangue , SolubilidadeRESUMO
The resonances of nonprotonated aromatic carbons in natural abundance 13C NMR spectra of hen egg white lysozyme are assigned to specific residues of the amino acid sequence. Chemical shift considerations, the effect of pH, and partially relaxed Fourier transform NMR spectra are used to assign each resonance to one of the seven types of nonprotonated aromatic carbons of amino acid residues. Spectra of chemically modified lysozyme samples yield various assignments to specific residues in the sequence. Line-broadening effects caused by binding of the relaxation probes Gd3+ and 4-N-acetamido-2,2,6,6-tetramethylipiperidine-1-oxyl yield specific assignments which are fully consistent with those based on chemical modifications. The effects of paramagnetic shift reagents and amino sugar inhibitors do not yield any obvious specific assignments. The effect of pH on the chemical shift of Cgamma of His-15 yields a pKalpha in agreement with published values, and indicates that the imidazole form of His-15 exists mainly (or entirely) as the Nepsilon3-H tautomer. The effect of pH on the chemical shifts (measured up to pH 8.8, at 38 degrees) of Czeta and Cgamma of the 3 tyrosine residues yields crude pKalpha values of 9.5 and 10 for Tyr-23 and one of the other tyrosines, respectively. The 3rd tyrosine residue does not exhibit titration behavior.