Do sequence repeats play an equivalent role in the choline-binding module of pneumococcal LytA amidase?

LytA amidase breaks down the N-acetylmuramoyl-L-alanine bonds in the peptid oglycan backbone of Streptococcus pneumoniae, Its polypeptide chain has two modules: the NH2-terminal module, responsible for the catalytic activity, and the COOH-terminal module, constructed by six tandem repeats of 20 or 21 amino acids (p1-p6) and a short COOH-terminal tail. The polypeptide chain must contain at least four repeats to efficiently anchor the autolysin to t he choline residues of the cell wall. Nevertheless, the catalytic efficienc y decreases by 90% upon deletion of the final tail. The structural implicat ions of deleting step by step the two last (p5 and p6) repeats and the fina l COOH-tail and their effects on choline-amidase interactions have been exa mined by comparing four truncated mutants with LytA amidase by means of dif ferent techniques. Removal of this region has minor effects on secondary st ructure content but significantly affects the stability of native conformat ions. The last 11 amino acids and the p5 repeat stabilize the COOH-terminal module; each increases the module transition temperature by about 6 degree s C, Moreover, the p5 motif also seems to participate, in a choline-depende nt way, in the stabilization of the NH2-terminal module. The effects of cho line binding on the thermal stability profile of the mutant lacking the p5 repeat might reflect a cooperative pathway providing molecular communicatio n between the choline-binding module and the NH2-terminal region, The three sequence motives favor the choline-amidase interaction, but the tail is an essential factor in the monomer <----> dimer self-association equilibrium of LytA and its regulation by choline, The final tail is required for prefe rential interaction of choline with LytA dimers and for the existence of di fferent sets of choline-binding sites. The p6 repeat scarcely affects the a midase stability but could provide the proper three dimensional orientation of the final tail.