H662 is the linchpin of ATP hydrolysis in the nucleotide-binding domain of the ABC transporter HlyB |
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| Authors: | Zaitseva Jelena Jenewein Stefan Jumpertz Thorsten Holland I Barry Schmitt Lutz |
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| Affiliation: | Institute of Biochemistry, Biocenter, Johann-Wolfgang Goethe University Frankfurt, Frankfurt, Germany. |
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| Abstract: | The ABC transporter HlyB is a central element of the HlyA secretion machinery, a paradigm of Type I secretion. Here, we describe the crystal structure of the HlyB-NBD (nucleotide-binding domain) with H662 replaced by Ala in complex with ATP/Mg2+. The dimer shows a composite architecture, in which two intact ATP molecules are bound at the interface of the Walker A motif and the C-loop, provided by the two monomers. ATPase measurements confirm that H662 is essential for activity. Based on these data, we propose a model in which E631 and H662, highly conserved among ABC transporters, form a catalytic dyad. Here, H662 acts as a 'linchpin', holding together all required parts of a complicated network of interactions between ATP, water molecules, Mg2+, and amino acids both in cis and trans, necessary for intermonomer communication. Based on biochemical experiments, we discuss the hypothesis that substrate-assisted catalysis, rather than general base catalysis might operate in ABC-ATPases. |
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| Keywords: | ATPase activity catalysis membrane transporter NBD-dimer X-ray crystallography |
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