Structure of Eznyme IIAlac from Lactococcus lactis - A Trimer of Three-Helical Bundles. Piotr Sliz1, Roswitha Engelmann2, Wolfgang Hengstenberg2, Emil F. Pai1 1Department of Biochemistry, University of Toronto, 1 King's College Circle, Toronto, ON M5S 1A8, Canada, 2Fakultat Biologie, Arbeitsgruppe Physiologie der Mikroorganismen, Ruhr-Universitat, Bochum, D-44780, Germany.
The phosphoenolpyruvate:sugar phosphotransferase system (PTS) is a complex, multiprotein bacterial system, which, through a cascade of several phosphorylation events, leads to the import and simultaneous phosphorylation of sugars. Along the phosphorylation cascade, the PTS is also involved in several regulatory events, acting as a primitive signal transduction system. The PTS consists of two general proteins, EI and HPr, and several sugar-specific enzymes II -- multiprotein permeases which are composed of two intracellular proteins, enzymes IIA and IIB, and a transmembrane channel, enzyme IIC.
We have determined the X-ray structure of enzyme IIA from the lactose- specific phosphoenolpyruvate transferase system (PTS) to a resolution of 2.3Å, using the SIRAS method. IIAlac is the first enzyme IIA of the lactose/cellobiose family for which the three-dimensional structure has been determined. The crystal structure reveals that although the motif of the phosphorylation site, two histidines, is preserved in IIAlac, its trimer of three helical bundles has no resemblance to the [beta] or a/[beta] topologies of other known enzymes IIA. The protein contains very interesting structural features, including a methionine cluster, a helical bundle which resembles a coiled-coil and a 230 Å3 hydrophobic cavity.
The lactose/cellobiose PTS family is the only one for which the high resolution X-ray structure has been determined for both of the intracellular components of the sugar-specific enzyme II, IIA and IIB. Since the other, general proteins of the PTS, EI and HPr are highly homologous between species, and the crystal structures of HPr and the phosphorylation domain of EI are known, the model of the phosphate transfer can now be studied in detail.