Structural characterization reveals a novel bilobed architecture for the ectodomains of insect stage expressed Trypanosoma brucei PSSA-2 and Trypanosoma congolense ISA.

African trypanosomiasis, caused by parasites of the genus Trypanosoma, is a complex of devastating vector-borne diseases of humans and livestock in sub-Saharan Africa. Central to the pathogenesis of African trypanosomes is their transmission by the arthropod vector, Glossina spp. (tsetse fly). Intriguingly, the efficiency of parasite transmission through the vector is reduced following depletion of Trypanosoma brucei Procyclic-Specific Surface Antigen-2 (TbPSSA-2). To investigate the underlying molecular mechanism of TbPSSA-2, we determined the crystal structures of its ectodomain and that of its homolog T. congolense Insect Stage Antigen (TcISA) to resolutions of 1.65 Å and 2.45 Å, respectively using single wavelength anomalous dispersion. Both proteins adopt a novel bilobed architecture with the individual lobes displaying rotational flexibility around the central tether that suggest a potential mechanism for coordinating a binding partner. In support of this hypothesis, electron density consistent with a bound peptide was observed in the inter-lob cleft of a TcISA monomer. These first reported structures of insect stage transmembrane proteins expressed by African trypanosomes provide potentially valuable insight into the interface between parasite and tsetse vector.

A comparison of the clavam biosynthetic gene clusters in Streptomyces antibioticus Tü1718 and Streptomyces clavuligerus.

The production of clavam metabolites has been studied previously in Streptomyces clavuligerus , a species that produces clavulanic acid as well as 4 other clavam compounds, but the late steps of the pathway leading to the specific end products are unclear. The present study compared the clavam biosynthetic gene cluster in Streptomyces antibioticus , chosen because it produces only 2 clavam metabolites and no clavulanic acid, with that of S. clavuligerus. A cosmid library of S. antibioticus genomic DNA was screened with a clavaminate synthase-specific probe based on the corresponding genes from S. clavuligerus, and 1 of the hybridizing cosmids was sequenced in full. A clavam gene cluster was identified that shows similarities to that of S. clavuligerus but also contains a number of novel genes. Knock-out mutation of the clavaminate synthase gene abolished clavam production in S. antibioticus, confirming the identity of the gene cluster. Knock-out mutation of a novel gene encoding an apparent oxidoreductase also abolished clavam production. A potential clavam biosynthetic pathway consistent with the genes in the cluster and the metabolites produced by S. antibioticus, and correspondingly different from that of S. clavuligerus, is proposed.