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Background: Cytochrome c is an integral part of the mitochondrial respiratory chain. It is confined to the intermembrane space of mitochondria, and has the function of transferring electrons between its redox partners. Solution studies of cytochrome c indicate that the conformation of the molecule is sensitive to the ionic strength of the medium., Results: The crystal structures of cytochromes c from several species have been solved at extremely high ionic strengths of near-saturated solutions of ammonium sulfate. Here we present the first crystal structure of ferricytochrome c at low ionic strength refined at 2.1 A resolution. In general, the structure has the same features as those determined earlier. However, there are some differences in both backbone and side-chain conformations in several areas. These areas coincide with those observed by NMR and resonance Raman spectroscopy to be sensitive to ionic strength., Conclusions: Neither ionic strength nor crystal-packing interactions have much influence on the conformation of horse cytochrome c. Nevertheless, some differences in the side-chain conformations at high and low ionic strengths may be important for understanding how the protein functions. Close examination of the gamma-turn (residues 27-29) conserved in cytochromes c leads us to propose the 'negative classical' gamma-turn to describe this unusual feature.

The inhibition of dihydrofolate reductase from chicken liver and from Lactobacillus casei has been studied with 4,6-diamino-1,2-dihydro-2,2-dimethyl-1-(substituted-phenyl)-s-triazines. It was found that for the chicken enzyme, inhibitor potency for 101 triazines was correlated by the following equation: log 1/Kiapp = 0.85 sigma tau' - 1.04 log (beta X 10 sigma tau' + 1) + 0.57 sigma + 6.36. The parameter tau' indicates that for certain substituents, tau = 0. In the case of the L. casei DHFR results, meta and para derivatives could not be included in the same equation. For 38 meta-substituted compounds, it was found that log 1/Kiapp = 0.38 tau'3-0.91 log (beta X 10 tau'3 + 1) + 0.71I + 4.60 and for 32 para-substituted phenyltriazines log 1/Kiapp = 0.44 tau'4-0.65 log (beta tau'4 + 1') - 0.90 upsilon + 0.69I + 4.67. In the L. casei equation, I is an indicator variable for substituents of the type CH2ZC6H4-Y and ZCH2C6H4-Y, where Z = O, NH, S, or Se. The parameter upsilon is Charton's steric parameter, which is similar to Taft's Es. The mathematical models obtained from correlation analysis are compared with stereo color graphics models.

In an X-ray diffraction study using the method of multiple isomorphous replacement, the structure of aspartate carbamoyltransferase (EC 2.1.3.2) complexed with the bisubstrate analog N-(phosphonacetyl)-L-aspartate (PALA) has been solved to 2.5 A. Ten rounds of model building and 123 cycles of restrained reciprocal space refinement have resulted in a model containing 94.4% of the theoretical atoms of the protein-inhibitor complex with an R-factor of 0.231. The fit of the model to the density is excellent, except for occasional side-chains and two sections of the regulatory chains that may be disordered. The electron density for the PALA molecule is readily identifiable for both catalytic (c) chains of the asymmetric unit and bonding interactions with several important residues including Ser52, Arg54, Thr55, Ser80, Lys84, Arg105, His134, Arg165, Arg229 and Gln231 are apparent. The carboxylate groups of the PALA molecule are in a nearly cis conformation. Gross quaternary changes between the T and R forms are noted and in agreement with earlier work from this laboratory. Namely, in the new structure the catalytic trimers move apart by 12 A along the 3-fold axis of the enzyme and relocate by 10 degrees relative to each other, adopting a more eclipsed position. The regulatory (r) chains in the new structure reorient about their 2-fold axis by 15 degrees. Large tertiary changes that include domain migration and rearrangement are also present between these two forms. In the R form both domains of the catalytic chain relocate closer to each other in order to bind to the inhibitor. The polar domain seems to bind primarily to the carbamoyl phosphate moiety of PALA, and the equatorial domain binds primarily to the L-aspartate moiety. Other changes in tertiary structure bring the 80s loop (from an adjacent catalytic chain) and the 240s loop into a position to interact with the PALA molecule. Changes have been searched for in all interface regions of the enzyme. While the C1-C4 and C1-R4 regions have been completely altered, most of the other interchain interfaces are similar in the T and R forms. The intrachain interfaces, between domains of the same catalytic chains, have undergone some reorganization as these domains move closer to each other when the inhibitor is bound. This new structure allows a reinterpretation of genetic and chemical modification studies done to date.(ABSTRACT TRUNCATED AT 400 WORDS)

In an x-ray diffraction study by the isomorphous replacement method, the structure of the complex of aspartate carbamoyltransferase (EC 2.1.3.2) bound to the bisubstrate analogue N-(phosphonacetyl)-L-aspartate has been solved to 2.9-A resolution (R = 0.24). The large quaternary structural changes previously deduced by molecular replacement methods have been confirmed: the two catalytic trimers (c3) move apart by 12 A and mutually reorient by 10 degrees, and the regulatory dimers (r2) reorient each about its twofold axis by about 15 degrees. In this, the T-to-R transition, new polar interactions develop between equatorial domains of c chains and the Zn domain of r chains. Within the c chain the two domains, one binding the phosphonate moiety (polar) and the other binding the aspartate moiety (equatorial) of the inhibitor N-(phosphonacetyl)-L-aspartate, move closer together. The Lys-84 loop makes a large relocation so that this residue and Ser-80 bind to the inhibitor of an adjacent catalytic chain within c3. A very large change in tertiary structure brings the 230-245 loop nearer the active site, allowing Arg-229 and Gln-231 to bind to the inhibitor. His-134 is close to the carbonyl group of the inhibitor, and Ser-52 is adjacent to its phosphonate group. However, no evidence exists in the literature for phosphorylation of serine in the mechanism. Residues studied by other methods, including Cys-47, Tyr-165, Lys-232, and Tyr-240, are too far from the inhibitor to have a direct interaction.