Mammalian alpha1,6-fucosyltransferase (FUT8) catalyses the transfer of a fucose residue from a donor substrate, guanosine 5'-diphosphate-beta-L-fucose to the reducing terminal N-acetylglucosamine (GlcNAc) of the core structure of an asparagine-linked oligosaccharide. Alpha1,6-fucosylation, also referred to as core fucosylation, plays an essential role in various pathophysiological events. Our group reported that FUT8 null mice showed severe growth retardation and emphysema-like lung-destruction as a result of the dysfunction of epidermal growth factor and transforming growth factor-beta receptors. To elucidate the molecular basis of FUT8 with respect to pathophysiology, the crystal structure of human FUT8 was determined at 2.6 A resolution. The overall structure of FUT8 was found to consist of three domains: an N-terminal coiled-coil domain, a catalytic domain, and a C-terminal SH3 domain. The catalytic region appears to be similar to GT-B glycosyltransferases rather than GT-A. The C-terminal part of the catalytic domain of FUT8 includes a Rossmann fold with three regions that are conserved in alpha1,6-, alpha1,2-, and protein O-fucosyltransferases. The SH3 domain of FUT8 is similar to other SH3 domain-containing proteins, although the significance of this domain remains to be elucidated. The present findings of FUT8 suggest that the conserved residues in the three conserved regions participate in the Rossmann fold and act as the donor binding site, or in catalysis, thus playing key roles in the fucose-transferring reaction.

Interleukin 15 (IL-15) and IL-2, which promote the survival of memory CD8(+) T cells and regulatory T cells, respectively, bind receptor complexes that share beta- and gamma-signaling subunits. Receptor specificity is provided by unique, nonsignaling alpha-subunits. Whereas IL-2 receptor-alpha (IL-2Ralpha) is expressed together in cis with the beta- and gamma-subunits on T cells and B cells, IL-15Ralpha is expressed in trans on antigen-presenting cells. Here we present a 1.85-A crystal structure of the human IL-15-IL-15Ralpha complex. The structure provides insight into the molecular basis of the specificity of cytokine recognition and emphasizes the importance of water in generating this very high-affinity complex. Despite very low IL-15-IL-2 sequence homology and distinct receptor architecture, the topologies of the IL-15-IL-15Ralpha and IL-2-IL-2Ralpha complexes are very similar. Our data raise the possibility that IL-2, like IL-15, might be capable of being presented in trans in the context of its unique receptor alpha-chain.

Lipoate-protein ligase A (LplA) catalyzes the formation of lipoyl-AMP from lipoate and ATP and then transfers the lipoyl moiety to a specific lysine residue on the acyltransferase subunit of ␣-ketoacid dehydrogenase complexes and on H-protein of the glycine cleavage system. The lypoyllysine arm plays a pivotal role in the complexes by shuttling the reaction intermediate and reducing equivalents between the active sites of the components of the complexes. We have determined the X-ray crystal structures of Escherichia coli LplA alone and in a complex with lipoic acid at 2.4 and 2.9 Å resolution, respectively. The structure of LplA consists of a large N-terminal domain and a small C-terminal domain. The structure identifies the substrate binding pocket at the interface between the two domains. Lipoic acid is bound in a hydrophobic cavity in the N-terminal domain through hydrophobic interactions and a weak hydrogen bond between carboxyl group of lipoic acid and the Ser-72 or Arg-140 residue of LplA. No large conformational change was observed in the main chain structure upon the binding of lipoic acid.Lipoic acid is a prosthetic group of acyltransferase (E2) subunit of the pyruvate, ␣-ketoglutarate, and branched-chain ␣-ketoacid dehydrogenase complexes and of H-protein of the glycine cleavage system (1-4). It attaches to a specific lysine residue on the proteins via an amide linkage between the ⑀-amino group of the lysine residue and the carboxyl group of lipoic acid. In the reaction sequence of the complexes, the lypoyllysine arm shuttles the reaction intermediates and reducing equivalents between the active sites of the components of the complexes.The attachment of lipoic acid to the proteins occurs by two-step reactions in which a lipoyl-AMP intermediate is formed from lipoic acid and ATP, and pyrophosphate is released in the initial activation reaction (Reaction 1).The lipoyl moiety of the intermediate is then transferred to apoproteins in the second transfer reaction, yielding the lipoylated protein and AMP (Reaction 2).Lipoyl-AMP ϩ apoprotein 3 lipoylated protein ϩ AMP (5, 6). LplA has a molecular mass of 37,795 Da, consisting of 337 amino acids excluding the initiating methionine residue, which is cleaved off during the biosynthesis (5). Strains with lplA null mutations have severe defects in the incorporation of exogenously supplied lipoic acid and lipoic acid analogues into apoproteins (5, 7). In E. coli, there is another enzyme, LipB, responsible for the covalent attachment of lipoic acid. LipB transfers lipoic acid/octanoic acid endogenously synthesized on the acyl carrier protein by the function of LipA to the lipoate-dependent enzymes (7-9). LipB consists of 213 amino acids, whose amino acid sequence shares only 12.7% identity with that of LplA (Fig. 1). On the other hand, the amino acid sequence of LplA shows 31 and 35% identity with those of human and bovine lipoyltransferase, the mammalian LplA homologues, respectively (Fig. 1). However, the mammalian lipoyltransferases have no ability to activate l...