J. Feeney

Suramin and suramin analogues inhibit merozoite surface protein-1 secondary processing and erythrocyte invasion by the malaria parasite Plasmodium falciparum

Malarial merozoites invade erythrocytes; and as an essential step in this invasion process, the 42-kDa fragment of Plasmodium falciparum merozoite surface protein-1 (MSP142) is further cleaved to a 33-kDa N-terminal polypeptide (MSP133) and an 19-kDa C-terminal fragment (MSP119) in a secondary processing step. Suramin was shown to inhibit both merozoite invasion and MSP142 proteolytic cleavage. This polysulfonated naphthylurea bound directly to recombinant P. falciparum MSP142 (Kd = 0.2 μm) and to Plasmodium vivax MSP142 (Kd = 0.3 μm) as measured by fluorescence enhancement in the presence of the protein and by isothermal titration calorimetry. Suramin bound only slightly less tightly to the P. vivax MSP133 (Kd = 1.5 μm) secondary processing product (fluorescence measurements), but very weakly to MSP119 (Kd ∼ 15 mm) (NMR measurements). Several residues in MSP119 were implicated in the interaction with suramin using NMR measurements. A series of symmetrical suramin analogues that differ in the number of aromatic rings and substitution patterns of the terminal naphthylamine groups was examined in invasion and processing assays. Two classes of analogue with either two or four bridging rings were found to be active in both assays, whereas two other classes without bridging rings were inactive. We propose that suramin and related compounds inhibit erythrocyte invasion by binding to MSP1 and by preventing its cleavage by the secondary processing protease. The results indicate that enzymatic events during invasion are suitable targets for drug development and validate the novel concept of an inhibitor binding to a macromolecular substrate to prevent its proteolysis by a protease.

A method of assigning 13C nuclear magnetic resonance spectra using europium(III) ion-induced pseudocontact shifts and C–H heteronuclear spin decoupling techniques

We report a novel method for making 13C n.m.r. assignments using lanthanide ion-induced pseudocontact shifts in the 1H spectrum to separate out the different absorptions and thus to facilitate the use of C–H selective heteronuclear spin decoupling experiments.

The 13C and 1H nuclear magnetic resonance spectra and methods of their assignment for nucleotides related to dihydronicotinamide adenine dinucleotide phosphate (NADPH)

The 13C n.m.r. spectra at 25·2 MHz of ribose-5-phosphate, AMP, NMN+, NAD+, NADH, NADP+, and NADPH have been assigned by use of a combination of techniques. In addition to established methods, such as comparisons of chemical shifts and coupling constants with those in model compounds, ionisation studies, and {1H}–13C heteronuclear selective decoupling experiments, we have described some novel methods of 13C spectral assignments, namely (i) addition of Eu3+ ions to induce pseudo-contact shifts in the proton spectrum to facilitate {1H}–13C decoupling experiments; (ii) the use of a graphical method of presenting results of off-resonance selective 1H irradiation experiments; (iii) paramagnetic shift predictions for 13C nuclei in non-rigid molecules in the presence of Eu3+ ions. Difficult proton assignments can sometimes be made on the basis of connecting assigned 13C signals with their corresponding proton resonance signals by heteronuclear decoupling.

Simplified 13C spectral assignments using a graphical method to present 13C spectra recorded under conditions of proton off-resonance spin decoupling

In the 13C spectrum of nicotinamide adenine dinucleotide (NAD+) the adenine and nicotinamide ring carbons are assigned by a graphical method of presenting the proton off-resonance decoupled 13C spectra as a function of proton irradiation frequency; this method is particularly applicable to closely spaced 13C signals.