Crystal structure of Thermotoga maritima 4-α-glucanotransferase and its acarbose complex: Implications for substrate specificity and catalysis

Anna Roujeinikova, Carsten Raasch, Svetlana Sedelnikova, Wolfgang Liebl, David W. Rice

Research output: Contribution to journalArticlepeer-review

72 Scopus citations


4-α-Glucanotransferase (GTase) is an essential enzyme in α-1,4-glucan metabolism in bacteria and plants. It catalyses the transfer of malto-oligosaccharides from an 1,4-α-D-glucan molecule to the 4-hydroxyl group of an acceptor sugar molecule. The crystal structures of Thermotoga maritima GTase and its complex with the inhibitor acarbose have been determined at 2.6 Å and 2.5 Å resolution, respectively. The GTase structure consists of three domains, an N-terminal domain with the (β/α)δ barrel topology (domain A), a 65 residue domain, domain B, inserted between strand β3 and helix α6 of the barrel, and a C-terminal domain, domain C, which forms an antiparallel β-structure. Analysis of the complex of GTase with acarbose has revealed the locations of five sugar-binding subsites (-2 to +3) in the active-site cleft lying between domain B and the C-terminal end of the (β/α)δ barrel. The structure of GTase closely resembles the family 13 glycoside hydrolases and conservation of key catalytic residues previously identified for this family is consistent with a double-displacement catalytic mechanism for this enzyme. A distinguishing feature of GTase is a pair of tryptophan residues, W131 and W218, which, upon the carbohydrate inhibitor binding, form a remarkable aromatic "clamp" that captures the sugar rings at the acceptor-binding sites +1 and +2. Analysis of the structure of the complex shows that sugar residues occupying subsites from -2 to +2 engage in extensive interactions with the protein, whereas the +3 glucosyl residue makes relatively few contacts with the enzyme. Thus, the structure suggests that four subsites, from -2 to +2, play the dominant role in enzyme-substrate recognition, consistent with the observation that the smallest donor for T. maritima GTase is maltotetraose, the smallest chain transferred is a maltosyl unit and that the smallest residual fragment after transfer is maltose. A close similarity between the structures of GTase and oligo-1,6-glucosidase has allowed the structural features that determine differences in substrate specificity of these two enzymes to be analysed.

Original languageEnglish
Pages (from-to)149-162
Number of pages14
JournalJournal of Molecular Biology
Issue number1
StatePublished - 2002
Externally publishedYes


  • 4-α-glucanotransferase
  • A-amylase family
  • Acarbose
  • Structure comparison
  • Thermotoga maritima


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