Parasite lactate dehydrogenase (pLDH) is a potential drug target for new antimalarials owing to parasite dependence on glycolysis for ATP production. The pLDH from all four species of human malarial parasites were cloned, expressed, and analyzed for structural and kinetic properties that might be exploited for drug development. pLDH from Plasmodium vivax, malariae, and ovale exhibit 90-92% identity to pLDH from Plasmodium falciparum. Catalytic residues are identical. Resides I250 and T246, conserved in most LDH, are replaced by proline in all pLDH. The pLDH contain the same five-amino acid insert (DKEWN) in the substrate specificity loops. Within the cofactor site, pLDH from P. falciparum and P. malariae are identical, while pLDH from P. vivax and P. ovale have one substitution. Homology modeling of pLDH from P. vivax, ovale, and malariae with the crystal structure of pLDH from P. falciparum gave nearly identical structures. Nevertheless, the kinetic properties and sensitivities to inhibitors targeted to the cofactor binding site differ significantly. Michaelis constants for pyruvate and lactate differ 8-9-fold; Michaelis constants for NADH, NAD(+), and the NAD(+) analogue 3-acetylpyridine adenine dinucleotide differ up to 4-fold. Dissociation constants for the inhibitors differ up to 21-fold. Molecular docking studies of the binding of the inhibitors to the cofactor sites of all four pLDH predict similar orientations, with the docked ligands positioned at the nicotinamide end of the cofactor site. pH studies indicate that inhibitor binding is independent of pH in the pH 6-8 range, suggesting that differences in dissociation constants for a specific inhibitor are not due to altered active site pK values among the four pLDH.

Triple-quadrupole mass spectrometry (MS/MS) has been used to compare nitroaromatic emissions from two different types of diesel engines, a direct-injection, single-cylinder engine and an indirect-injection V-8 engine. The low level of nitropyrenes/ nitrofluoranthenes in exhaust extracts may be due in part to the low pyrene content of the reference fuel used. Addition of pyrene to reference fuel resulted in increased pyrene in exhaust extracts, but only minor differences in mutagenicity. Only about 1/1000 of the total mutagenicity from complete reaction with N02 appeared to have occurred during diesel exhaust and soot collection on filters. Fractionation with Me2SO was found to be useful in separating aliphatic hydrocarbons from mutagenic activities and in concentrating nitroaromatic compounds for MS/MS analysis.Concentration of certain nitroaromatic compounds was necessary for isobutane chemical ionization MS/MS, while atmospheric pressure MS/MS appeared capable of detecting nitroaromatic compounds even in unfractionated extracts. MS/MS comparisons of concentrated samples of differing mutagenicities showed the main differences were increased ion intensities of dinitro compounds in more mutagenic samples. It is concluded that the polynitro compounds may be of more significance than mononitro compounds in the mutagenic activities that have been found associated with diesel soot.