35 years of resistance

The misuse of antibiotics is not only a medical problem, but also a major agricultural one. The US Food and Drug Administration has recently shifted its focus to voluntary oversight.     

R-state haemoglobin with low oxygen affinity: crystal structures of deoxy human and carbonmonoxy horse haemoglobin bound to the effector molecule L35

Although detailed crystal structures of haemoglobin (Hb) provide a clear understanding of the basic allosteric mechanism of the protein, and how this in turn controls oxygen affinity, recent experiments with artificial effector molecules have shown a far greater control of oxygen binding than with natural heterotropic effectors. Contrary to the established text-book view, these non-physiological compounds are able to reduce oxygen affinity very strongly without switching the protein to the T (tense) state. In an earlier paper we showed that bezafibrate (BZF) binds to a surface pocket on the alpha subunits of R state Hb, strongly reducing the oxygen affinity of this protein conformation. Here we report the crystallisation of Hb with L35, a related compound, and show that this binds to the central cavity of both R and T state Hb. The mechanism by which L35 reduces oxygen affinity is discussed, in relation to spectroscopic studies of effector binding.     

Cloning of schistosomes

Before cloning was perfected, the only way to maintain schistosome strains was by sustaining the entire parasite cycle, ie. by ensuring the regular transmission of the parasite from the snail to the vertebrate, then from the vertebrate to the snail, and so on ad infinitum. Joseph Jourdane explains that by cloning the parasite in the snail and transplanting the larval parasite from snail to snail, the passage in vertebrates can be avoided indefinitely.     

Effect of ionizing radiation on haemoglobin: the oxy-derivative of haemoglobin Iwate

It is well established that exposure of oxyhaemoglobin to ionizing radiation results in remarkably selective electron addition to the (FeO2) unit, giving a novel species, (FeO2)-, in which the extra electron is largely localized on iron and dioxygen. This work has now been extended to haemoglobin (Hb.) Iwate. The haemoglobin M. Iwate used is a mutant haemoglobin having only Fe(III) alpha-chains by oxy beta-chains (alpha 2 Met beta 2 oxy). The haem iron atoms in the alpha-chains are coordinated in the fifth site by a proximal tyrosine in place of histidine. This unit is a high-spin Fe(III) with axial symmetry and prominent electron spin resonance (ESR) features in the g = 6 and g = 2 regions. On exposure to 60Co gamma-rays at 77 K, efficient electron addition occurred at both types of iron centre, giving Fe(II) and (FeO2)- units. The former was monitored by the decrease of the g = 6 feature for Fe(III) and the latter by the growth of g-features at 2.254 (gx), 2.149 (gy) and 1.967 (gz). These values are close to those for the FeO2- centre formed in the beta-chains of normal oxyhaemoglobin. On annealing above 77 K, two changes occurred: first there was a small but clear increase in gx and gy, followed by a marked reduction in gx and gy giving g-values close to those for the centre formed directly in the alpha-chains of the normal protein. Finally, this intermediate species gave a centre having gx = 2.310, gy = 2.180 and gz = 1.935. These values are typical of low-spin Fe(III) haemoglobin and are assigned to the protonated complex, Fe(III)O2H. Ultimately at ca. room temperature, this was converted into the high-spin, met-form, with a gain in the g = 6 feature. These results established that the beta-chain centre in Hb. Iwate behave in the same way as isolated beta-chains. They also confirm that electron addition to the oxy-units is facile, even in the presence of Fe(III) units in each tetramer. The results also confirm that electron capture to give (FeO2)- units is not followed by internal electron-transfer to give Fe(II) from the Fe(III) centres in the alpha-chains.     

Virulence factors of schistosomes

This review considers whether the products of schistosomes in the mammalian host can be considered as virulence factors. These include: the cercarial secretions used in infection, those of the migrating schistosomulum, surface-exposed proteins of adult worms in the portal system and their gut vomitus in the context of immune evasion, secretions of the egg facilitating its escape from gut tissues and micro-exon gene products.     

Genetic manipulation of schistosomes

In contrast to the situations with model organisms and parasitic protozoa, progress with gene manipulation with schistosomes has been delayed by impediments that include our inability to maintain the life cycle in vitro, absence of immortalized cell lines, large genome sizes, unavailability of drug resistance markers and other difficulties. However, in the past few years, tangible progress has been reported towards development of tools for gene manipulation and transgenesis of schistosomes, and there is reason to believe that the field is on the verge of transformation into an era where genetic manipulation is routine. Recent reports dealing with approaches and tools to manipulate the genome and gene expression in schistosomes are reviewed here.     

Phenol oxidase activity: inductionin female schistosomes by in vitro incubation.

A biochemical methods has been developed for detecting phenol oxidase in female Schistosoma mansoni. Enzyme activity is observed only after incubation of the female schistosomes for an extended period of time in tissue culture media. Male S. mansoni do not contain detectable levels of phenol oxidase activity. The properties of this enzyme are similar to those identified for a phenol oxidase from Fasciola hepatica. L-DOPA, dopamine, and tyrosine were found to be good substrates for this enzyme. Vmax = 14.1, 8.1, and 6.1 mumoles O2/min/mg protein for each substrate, respectively. This enzyme appears to be associated with egg production and thus may be a useful marker for biochemical and immunological studies.