PDBsum entry 1n4a

Transport protein

PDB id

1n4a

Contents

			Protein chains

					244 a.a. *

			Ligands

					CNC ×2

			Waters ×549

* Residue conservation analysis

References listed in PDB file

Key reference

Title

Crystal structures of the btuf periplasmic-Binding protein for vitamin b12 suggest a functionally important reduction in protein mobility upon ligand binding.

Authors

N.K.Karpowich, H.H.Huang, P.C.Smith, J.F.Hunt.

Ref.

J Biol Chem, 2003, 278, 8429-8434. [DOI no: 10.1074/jbc.M212239200]

PubMed id

12468528

Abstract

BtuF is the periplasmic binding protein (PBP) for the vitamin B12 transporter BtuCD, a member of the ATP-binding cassette (ABC) transporter superfamily of transmembrane pumps. We have determined crystal structures of Escherichia coli BtuF in the apo state at 3.0 A resolution and with vitamin B12 bound at 2.0 A resolution. The structure of BtuF is similar to that of the FhuD and TroA PBPs and is composed of two alpha/beta domains linked by a rigid alpha-helix. B12 is bound in the "base-on" or vitamin conformation in a wide acidic cleft located between these domains. The C-terminal domain shares structural homology to a B12-binding domain found in a variety of enzymes. The same surface of this domain interacts with opposite surfaces of B12 when comparing ligand-bound structures of BtuF and the homologous enzymes, a change that is probably caused by the obstruction of the face that typically interacts with this domain by the base-on conformation of vitamin B12 bound to BtuF. There is no apparent pseudo-symmetry in the surface properties of the BtuF domains flanking its B12 binding site even though the presumed transport site in the previously reported crystal structure of BtuCD is located in an intersubunit interface with 2-fold symmetry. Unwinding of an alpha-helix in the C-terminal domain of BtuF appears to be part of conformational change involving a general increase in the mobility of this domain in the apo structure compared with the B12-bound structure. As this helix is located on the surface likely to interact with BtuC, unwinding of the helix upon binding to BtuC could play a role in triggering release of B12 into the transport cavity. Furthermore, the high mobility of this domain in free BtuF could provide an entropic driving force for the subsequent release of BtuF required to complete the transport cycle.



	Figure 2. Fig. 2. Structural homologues of BtuF. A, stereo pair of the structural alignment of the N-terminal domains of BtuF and E. coli FhuD bound to gallichrome (PBD accession number 1EFD). BtuF is colored by domain as above, and vitamin B12 is shown in ball-and-stick representation and colored magenta. FhuD is colored gold with the bound gallichrome shown in bright green. B, stereo pair of the structural alignment of domain II of BtuF with the B12-binding domain of E. coli methionine synthase (MetE, PDB accession number 1BMT). The corrin ring and DMB of B12 are colored brown and red, respectively. MetE is colored blue with the corrin ring and DMB of its bound B12 colored aqua and cyan, respectively.		Figure 3. Fig. 3. Structure of the Vitamin B12 binding site of BtuF. A, stereo pair of the B12-binding site of BtuF colored as in Fig. 1A, with conserved waters represented as green spheres. The side chains of relevant residues are depicted in ball-and-stick representation colored by domain with associated nitrogen and oxygen atoms blue and red, respectively. Direct and water-mediated H-bonds are represented by red and green dotted lines, respectively. B, surface representation of the likely BtuCD-interacting face of BtuF color-ramped according to sequence conservation, with white indicating no conservation and burgundy indicating 100% conservation in the five known BtuFs. C, surface representation of the same face of BtuF color-ramped according to electrostatic potential, with red indicating negative potential, blue indicating positive potential, and fully saturated colors indicating potential ± 5 kT (assuming an ionic strength of 100 mM).

PROCHECK

	Headers