The structure of rhamnose isomerase from Escherichia coli and its relationwith xylose isomerase illustrates a change between inter and intra-subunitcomplementation during evolution

Using a new expression construct, rhamnose isomerase from Escherichia coli was purified and crystallized. The crystal structure was solved by multiple isomorphous replacement and refined to a crystallographic residual of 17.4 % at 1.6 Angstrom resolution. Rhamnose isomerase is a tight tetramer of fo ur (beta/alpha)(8)-barrels. A comparison with other known structures reveal s that rhamnose isomerase is most similar to xylose isomerase. Alignment of the sequences of the two enzymes based on their structures reveals a hithe rto undetected sequence identity of 13 %, suggesting that the two enzymes e volved from a common precursor. The structure and arrangement of the (beta/ alpha)(8)-barrels of rhamnose isomerase are very similar to xylose isomeras e. Each enzyme does, however, have additional a-helical domains, which are involved in tetramer association, and largely differ in structure. The stru ctures of complexes of rhamnose isomerase with the inhibitor L-rhamnitol an d the natural substrate L-rhamnose were determined and suggest that an exte nded loop, which is disordered in the native enzyme, becomes ordered on sub strate binding, and may exclude bulk solvent during catalysis. Unlike xylos e isomerase, this loop does not extend across a subunit interface but contr ibutes to the active site of its own subunit. It illustrates how an interco nversion between inter and intra-subunit complementation can occur during e volution. In the crystal structure (although not necessarily in vivo) rhamn ose isomerase appears to bind Zn2+ at a "structural" site. In the presence of substrate the enzyme also binds Mn2+ at a nearby "catalytic" site. An ar ray of hydrophobic residues, not present in xylose isomerase, is Likely to be responsible for the recognition of L-rhamnose as a substrate. The availa ble structural data suggest that a metal-mediated hydride-shift mechanism, which is generally favored for xylose isomerase, is also feasible for rhamn ose isomerase. (C) 2000 Academic Press.