5KC0

Crystal structure of TmRibU, hexagonal crystal form

Summary for 5KC0

• Entry DOI: 10.2210/pdb5kc0/pdb
• Descriptor: Riboflavin transporter RibU, nonyl beta-D-glucopyranoside, RIBOFLAVIN (3 entities in total)
• Functional Keywords: membrane transport, integral membrane protein, vitamins, thermophile, transport protein-membrane protein complex, transport protein/membrane protein
• Biological source: Thermotoga maritima (strain ATCC 43589 / MSB8 / DSM 3109 / JCM 10099)
• Cellular location: Cell inner membrane ; Multi-pass membrane protein : Q9X1G6
• Total number of polymer chains: 1
• Total formula weight: 21775.00

Authors

Karpowich, N.K.,Wang, D.N.,Song, J.M. (deposition date: 2016-06-03, release date: 2016-06-29, Last modification date: 2024-12-25)

Primary citation

Karpowich, N.K.,Song, J.,Wang, D.N.
An Aromatic Cap Seals the Substrate Binding Site in an ECF-Type S Subunit for Riboflavin.
J.Mol.Biol., 428:3118-3130, 2016

PubMed Abstract: ECF transporters are a family of active membrane transporters for essential micronutrients, such as vitamins and trace metals. Found exclusively in archaea and bacteria, these transporters are composed of four subunits: an integral membrane substrate-binding subunit (EcfS), a transmembrane coupling subunit (EcfT), and two ATP-binding cassette ATPases (EcfA and EcfA'). We have characterized the structural basis of substrate binding by the EcfS subunit for riboflavin from Thermotoga maritima, TmRibU. TmRibU binds riboflavin with high affinity, and the protein-substrate complex is exceptionally stable in solution. The crystal structure of riboflavin-bound TmRibU reveals an electronegative binding pocket at the extracellular surface in which the substrate is completely buried. Analysis of the intermolecular contacts indicates that nearly every available substrate hydrogen bond is satisfied. A conserved aromatic residue at the extracellular end of TM5, Tyr130, caps the binding site to generate a substrate-bound, occluded state, and non-conservative mutation of Tyr130 reduces the stability of this conformation. Using a novel fluorescence binding assay, we find that an aromatic residue at this position is essential for high-affinity substrate binding. Comparison with other S subunit structures suggests that TM5 and Loop5-6 contain a dynamic, conserved motif that plays a key role in gating substrate entry and release by S subunits of ECF transporters.

PubMed: 27312125
DOI: 10.1016/j.jmb.2016.06.003

Experimental method

X-RAY DIFFRACTION (3.2001 Å)