(2) Proton release to extracellular side
 

(3)Retinal curvature change and connectivity change of the retinal Schiff base
 

(4) Protein conformational change and proton uptake from cytoplasm
 

A classification by Saier and colleagues1 (www-biology.ucsd.edu/~msaier/transport) groups membrane transporters into four categories: channels and pores, pumps, porters, and group translocators. Together, these proteins are thought to make up as much as 30% of all integral membrane proteins, with a majority contributed by molecules that function as pumps and porters. Significant progress has been made towards the structural analysis of proteins that function as channels, pores and pumps, but no atomic structures are available yet for any member of the porter family. Of the 107 recognized families of porters, the Major Facilitator Superfamily represents the largest grouping (37 families), and is thought to include over a thousand evolutionarily related members whose functions range from the transport of metabolites and antibiotics in bacteria to neurotransmitter transport in human synaptic membranes. Representative examples of the MFS include the bacterial lactose permease (LacY) and tetracycline transporter (Tet A(B)) and the human glucose transporter (GLUT1). We have recently reported the three-dimensional structure of a 44 kDa bacterial representative of this superfamily, OxlT, which carries out the selective exchange of oxalate for formate across the cytoplasmic membrane of the bacterium Oxalobacter formigenes.

Stereo view of the structure of the oxalate transporter at 6.5 Å resolution
(see Hirai et al Nat. Struct. Biol. 9, 597-600 (2002).  

Charge potential distribution in the cytoplasmically open state of OxlT as viewed from the cytoplasmic side, with blue and red colors denoting positive and negative potentials respectively. The map was prepared based on the structural model derived for OxlT by electron crystallography. The location of the band of positive potential in the center of the channel provides a structural explanation for attraction of the negatively charged oxalate substrate.
 

For more details, see Hirai and Subramaniam (2004).
 

(ii) Molecular structure of the pyruvate dehydrogenase complex.

Understanding how multi-component molecular machines function and how multi-step reactions are catalysed is an emerging frontier in cell biology, which will begin to define the black box that exists between our knowledge of the structures of individual proteins and those of cellular organelles. We have determined the three-dimensional structures at ~ 27 Å resolution for two giant icosahedral pyruvate dehydrogenase complexes from B. stearothermophilus using electron cryo-microscopy: one is a 11 MDa complex composed of 60 copies of E1 and E2 enzymes, and another is a 9 MDa complex composed of 60 copies of E2 and E3 enzymes. By positioning the previously determined structures of E1, E3 and the three domains of E2 into the model, we have arrived at atomic interpretations for the entire E1E2 and E2E3 complexes. To our knowledge, these are the largest non-viral protein complexes for which such atomic models are available, and they provide unique insights into the functional mechanism of a fascinating cellular machine that remains inaccessible to structural analysis by X-ray crystallography.

For more details see Milne et al EMBO J. (2002) and Milne et al (2006).

Architecture, at 27 Å resolution, of pyruvate dehydrogenase, and proposed mechanism for
synthesis of acetyl CoA derived by electron microscopy.