The crystal structure of a liganded trehalose/maltose-binding protein fromthe hyperthermophilic Archaeon Thermococcus litoralis at 1.85 angstrom

We report the crystallization and structure determination at 1.85 Angstrom of the extracellular, membrane-anchored trehalose/maltose-binding protein ( TMBP) in complex with its substrate trehalose. TMBP is the substrate recogn ition site of the high-affinity trehalose/maltose ABC transporter of the hy perthermophilic Archaeon Thermococcus litoralis. In vivo, this protein is a nchored to the membrane, presumably via an N-terminal cysteine lipid modifi cation. The crystallized protein was N-terminally truncated, resulting in a soluble protein exhibiting the same binding characteristics as the wild-ty pe protein. The protein shows the characteristic features of a transport-re lated, substrate-binding protein and is structurally related to the maltose -binding protein (MBP) of Escherichia coli. It consists of two similar lobe s, each formed by a parallel beta -sheet flanked by alpha -helices on both sides. Both are connected by a hinge region consisting of two anti parallel beta -strands and an alpha -helix. As in MBP, the substrate is bound in th e cleft between the lobes by hydrogen bonds and hydrophobic interactions. H owever, compared to maltose binding in MBP, direct hydrogen bonding between the substrate and the protein prevails while apolar contacts are reduced. To elucidate factors contributing to thermostability, we compared TMBP with its mesophilic counterpart MBP and found differences known from similar in vestigations. Specifically, we find helices that are longer than their stru cturally equivalent counterparts, and fewer internal cavities. (C) 2001 Aca demic Press.