The structure of enzyme IIAlactose from Lactococcus lactis reveals a new fold and points to possible interactions of a multicomponent system.
BACKGROUND
The bacterial phosphoenolpyruvate: sugar phosphotransferase system (PTS) is responsible for the binding, transmembrane transport and phosphorylation of numerous sugar substrates. The system is also involved in the regulation of a variety of metabolic and transcriptional processes. The PTS consists of two non-specific energy coupling components, enzyme I and a heat stable phosphocarrier protein (HPr), as well as several sugar-specific multiprotein permeases known as enzymes II. In most cases, enzymes IIA and IIB are located in the cytoplasm, while enzyme IIC acts as a membrane channel. Enzyme IIAlactose belongs to the lactose/cellobiose-specific family of enzymes II, one of four functionally and structurally distinct groups. The protein, which normally functions as a trimer, is believed to separate into its subunits after phosphorylation.


RESULTS
The crystal structure of the trimeric enzyme IIAlactose from Lactococcus lactis has been determined at 2.3 A resolution. The subunits of the enzyme, related to each other by the inherent threefold rotational symmetry, possess interesting structural features such as coiled-coil-like packing and a methionine cluster. The subunits each comprise three helices (I, II and III) and pack against each other forming a nine-helix bundle. This helical bundle is stabilized by a centrally located metal ion and also encloses a hydrophobic cavity. The three phosphorylation sites (His78 on each monomer) are located in helices III and their sidechains protrude into a large groove between helices I and II of the neighbouring subunits. A model of the complex between phosphorylated HPr and enzyme IIAlactose has been constructed.


CONCLUSIONS
Enzyme IIAlactose is the first representative of the family of lactose/cellobiose-specific enzymes IIA for which a three-dimensional structure has been determined. Some of its structural features, like the presence of two histidine residues at the active site, seem to be common to all enzymes no overall structural homology is observed to any PTS proteins or to any other proteins in the Protein Data Bank. Enzyme IIAlactose shows surface complementarity to the phosphorylated form of HPr and several energetically favourable interactions between the two molecules can be predicted.
Sliz
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Department of Biochemistry, University of Toronto, ON, Canada.
Engelmann
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Hengstenberg
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Pai
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EF
D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D018360:Crystallography, X-Ray; D013294:Lactococcus lactis / Q000201:enzymology*; D007785:Lactose / Q000378:metabolism*; D008670:Metals / Q000737:chemistry / Q000378:metabolism; D008715:Methionine; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System / Q000737:chemistry* / Q000378:metabolism*; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D013211:Staphylococcus aureus / Q000201:enzymology
D001426:Bacterial Proteins; D008670:Metals; D008715:Methionine; D010731:Phosphoenolpyruvate Sugar Phosphotransferase System; C030182:phosphocarrier protein HPr; D007785:Lactose