The predominant orientation of the phosphorylcholine polar head group in phosphatidylcholine and sphingomyelin bilayers and cholesterol perturbations of that orientation have been identified by exploiting the 31P (1H) nuclear Overhauser effect (NOE) in the 31P NMR spectra of phospholipid bilayers. In pure egg phosphatidylcholine bilayers, a NOE of 40% is observed. The magnitude of the NOE has been measured as a function of continuous-wave proton-decoupler frequency in order to identify the proton source of the NOE. In pure egg phosphatidylcholine bilayers, the maximum NOE occurs at the N-methyl proton resonance position of the choline moiety. In a modified phosphatidylcholine in which all the N-methyl protons were replaced by deuterium, the NOE arose from methylene protons next to the phosphate. In mixed systems of phosphatidylcholine and phosphatidylethanolamine, and phosphatidylcholine and diphosphatidylglycerol, both phospholipid resonances attained maximum NOE at the position of the N-methyl proton resonance of phosphatidylcholine. An analogous result was obtained with pure sphingomyelin. These results are explained by orienting the phosphorylcholine portion of the molecule parallel to the surface of the bilayer so that the positively charged N-methyl moiety is located close to the negatively charged phosphate on a neighboring phospholipid in an intermolecular interaction. Addition of cholesterol is shown to disrupt the intermolecular interaction in phosphatidylcholine bilayers.

Rates of N-demethylation of N,N-dimethylaniline and of eight meta- or para-substituted N,N-dimethylanilines by rat liver cytochrome P-450PB-B (P-450) were determined under conditions where oxidation was supported by iodosylbenzene or NADPH-P-450 reductase. The rates of dimethylaniline oxidation were found to correlate with the substrate oxidation-reduction potential within each series of substrates supported by a particular oxygen activation protocol; the kcat for each substrate studied was approximately 20-fold faster in the iodosylbenzene-supported system relative to the NADPH-P-450 reductase supported system. Since the N-demethylation of amines is believed to proceed via an initial electron-transfer step, a kinetic scheme for P-450 was proposed that enabled evaluation of the data according to theoretical treatments that correlate rates of electron transfer with extrakinetic parameters. In these analyses, the data could be fitted to the Rehm-Weller and Agmon-Levine equations, providing lambda values (for the energetics of enzyme-substrate reorganization) of 22-26 kcal mol-1 and apparent E1/2 (oxidation-reduction potentials) of 1.7-2.0 V (vs saturated calomel) for the oxidized enzyme. The apparent E1/2 for the enzyme is composed of contributions from the intrinsic potential of the active prosthetic core of the enzyme, the Fe = O - porphyrin species, and a coulombic factor that is a function of the charge-separated radical anion/radical cation pair produced upon electron transfer.(ABSTRACT TRUNCATED AT 250 WORDS)

Of the two Ca(II) in carp muscle calcium binding parvalbumin B, one may be removed by dialysis against EGTA without significant alteration of the circular dichroism spectra at 224 nm, a result suggesting little or no change in the helical content of 47%. Binding of the two Ca(II) is not cooperative. Addition of a 20-fold or greater excess of EGTA at pH 8.5 results in removal of both Ca(II), reduces the helical content to about 39%, alters only served in Tris buffer at pH 8.5 or bis-tris buffer at pH 6.5.1 Abbreviations used are: Nbs2, 5,5'-dithiobis(2-nitrobenzoic acid); EGTA, ethylene glycol bis(/3-aminoethyl ether)-7V,yV'-tetraacetate. BIOCHEMISTRY,VOL.

In order to obtain information about aluminum(III)-phosphate interactions, potentiometric measurements were carried out to characterize the complex forming properties of Al(III) with organic phosphates, phosphonates, and nucleoside-5'-monophosphates. The aluminum(III)-orthophosphate system is difficult to study due to AlPO(4) precipitation. To overcome this problem, the stability constant logarithms of the 1:1 Al(III) complexes of ligands with the same donor groups (log K(1:1)) were plotted against the basicities of the ligands (log K(PO)3(H)). The resulting linear free energy relation (LFER) indicates that organic phosphates, phosphonates, and uridine-, thymidine-, and guanosine 5'-monophosphates similarly bind Al(III). Adenosine and cytidine 5'-monophosphate fall above the LFER owing to the presence of a second microform with the nucleic base protonated and a hydroxide bound to the Al(III). From the LFER the log stability constant for Al(III) binding to HPO(4)(2-) is estimated as 6.13 +/- 0.05. From the weakness of any soluble orthophosphate complexes of Al(III) we confirm the importance of citrate as the main small molecule Al(3+) binder in the blood serum. The study includes investigation of Al(III) binding to di- and triphosphates, which bind metal ion differently than monophosphates. Structures of the complexes were supported by (31)P NMR measurements.

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