Structural analysis of adenylate cyclases from Trypanosoma brucei in theirmonomeric state

Cyclic AMP is a major trigger of the differentiation process of Trypanosoma brucei, a bloodstream parasite causing sleeping sickness. Its generation i n trypanosomes is accomplished by a unique battery of membrane-bound adenyl ate cyclases (ACs). We have determined the high-resolution X-ray structures of the catalytic domains of two trypanosomal ACs (tACs), GRESAGL4.1 and GR ESAG4.3. The tAC domains are structurally highly related to the AC domains of higher eukaryotes, but also comprise a highly conserved structural eleme nt near the active site, the Delta -subdomain. A cavity below the Delta -su bdomain might correspond to an allosteric regulator site as indicated by th e stereospecific binding of a single (2S,3S)-1,4-dimercapto-2,3-butanediol molecule. In three different crystal forms, the tAC domains are exclusively observed in a monomeric, catalytically inactive state. Biochemical analysi s and the mutagenesis profile of GRESAG4.1 confirmed a common catalytic mec hanism of tACs that involves transient dimerization of the AC domain. A low dimerization tendency might play a regulatory role in T.brucei if the acti vation of tACs is similarly driven by ligand-induced dimerization as in mem brane-bound guanylate cyclases.