PDBsum entry: 1f0p

Transferase

PDB id: 1f0p

Contents

Protein chain

284 a.a. *

Ligands

MES

TRE ×2

MPD ×3

Waters ×223

* Residue conservation analysis

PDB id:

1f0p

Name:

Transferase

Title:

Mycobacterium tuberculosis antigen 85b with trehalose

Structure:

Antigen 85-b. Chain: a. Engineered: yes

Source:

Mycobacterium tuberculosis. Organism_taxid: 1773. Expressed in: mycobacterium smegmatis. Expression_system_taxid: 1772.

Resolution:

1.90Å R-factor: 0.195 R-free: 0.284

Authors:

D.H.Anderson,G.Harth,M.A.Horwitz,D.Eisenberg,Tb Structural Genomics Consortium (Tbsgc)

Key ref:

D.H.Anderson et al. (2001). An interfacial mechanism and a class of inhibitors inferred from two crystal structures of the Mycobacterium tuberculosis 30 kDa major secretory protein (Antigen 85B), a mycolyl transferase. J Mol Biol, 307, 671-681. PubMed id: 11254389 DOI: 10.1006/jmbi.2001.4461

Date:

16-May-00 Release date: 24-Jan-01

Protein chain

P0C5B9  (A85B_MYCTU) -  Antigen 85-B

Seq: 325 a.a.

Struc: 284 a.a.

 Gene Ontology (GO) functional annotation 

• Cellular component: extracellular region (3 terms)
• Biological process: growth (4 terms)
• Biochemical function: protein binding (4 terms)

DOI no: 10.1006/jmbi.2001.4461

J Mol Biol 307:671-681 (2001)

PubMed id: 11254389

An interfacial mechanism and a class of inhibitors inferred from two crystal structures of the Mycobacterium tuberculosis 30 kDa major secretory protein (Antigen 85B), a mycolyl transferase.

D.H.Anderson, G.Harth, M.A.Horwitz, D.Eisenberg.

ABSTRACT

The Mycobacterium tuberculosis 30 kDa major secretory protein (antigen 85B) is the most abundant protein exported by M. tuberculosis, as well as a potent immunoprotective antigen and a leading drug target. A mycolyl transferase of 285 residues, it is closely related to two other mycolyl transferases, each of molecular mass 32 kDa: antigen 85A and antigen 85C. All three catalyze transfer of the fatty acid mycolate from one trehalose monomycolate to another, resulting in trehalose dimycolate and free trehalose, thus helping to build the bacterial cell wall. We have determined two crystal structures of M. tuberculosis antigen 85B (ag85B), initially by molecular replacement using antigen 85C as a probe. The apo ag85B model is refined against 1.8 A data, to an R-factor of 0.196 (R(free) is 0.276), and includes all residues except the N-terminal Phe. The active site immobilizes a molecule of the cryoprotectant 2-methyl-2,4-pentanediol. Crystal growth with addition of trehalose resulted in a second ag85B crystal structure (1.9 A resolution; R-factor is 0.195; R(free) is 0.285). Trehalose binds in two sites at opposite ends of the active-site cleft. In our proposed mechanism model, the trehalose at the active site Ser126 represents the trehalose liberated by temporary esterification of Ser126, while the other trehalose represents the incoming trehalose monomycolate just prior to swinging over to the first trehalose site to displace the mycolate from its serine ester. Our proposed interfacial mechanism minimizes aqueous exposure of the apolar mycolates. Based on the trehalose-bound structure, we suggest a new class of antituberculous drugs, made by connecting two trehalose molecules by an amphipathic linker.

Selected figure(s)

Figure 1.
Figure 1. Ribbon diagram of trehalose-bound ag85B, showing its α-β hydrolase fold. Molscript [Kraulis 1991] automatically assigned the secondary structural elements by its backbone φ and ψ criteria, and produced most of this Figure. Cyan ribbons represent α-helices, magenta ribbons β-strands, coil and turn regions are white. The N terminus is at upper left, and the C terminus is at the back of the molecule. Active site Ser126 and His262 CA and side-chain atoms are shown (protein C atoms are gray; N, blue; O, red). Trehalose 1151 is on the left, Trehalose 1152 is on the far right, and MPD 1001 is at the center (all with yellow C atoms and red O atoms). The MPD at this site may be a surrogate for the mycolate ester intermediate. Figure 1, Figure 3 and Figure 5 were produced using XtalView [McRee 1993], and were rendered with Raster3D [Merritt and Bacon 1997].

Figure 2.
Figure 2. Schematic diagrams of trehalose hydrogen bonds to ag85B protein. Hydrogen bond contacts were determined automatically with XtalView [McRee 1993]. (a) Trehalose 1151 binds at the active site Ser126, has many contacts, and is well ordered in the crystal. Wat401 was the most prominent non-protein density in the apo ag85B maps. Nearby hydrophobic atoms are: Leu42 CB, Pro225 CB, Leu229 CD1, catalytic triad His262 CD2 and Trp267 CH2 and CZ3. Trehalose binding at this site displaced five water molecules from the apo ag85B structure: water molecules 418, 442, 455, 528, 574. In addition, Wat567 binds to Arg43 in the apo structure in a site not overlapping Tre1151, but is absent from the trehalose model. Water molecules 595, 603 and 615 bind along with Tre1151. (b) Trehalose 1152 binds at the other end of the active site groove. In the text and Figure 4 we propose that this second site is the second trehalose monomycolate substrate-binding site. Fewer protein contacts result in more disorder and worse density for the upper (primed) ring (Figure 3(c). Nearby hydrophobic atoms are: Met159 CE (disordered in apo ag85B), Phe232 ring, CD1 atoms of leucine residues 152 and 163. Binding trehalose at the non-catalytic Tre1152 site displaces water molecules 460 and 543 of the apo ag85B structure, and adds the new Wat567.

The above figures are reprinted by permission from Elsevier: J Mol Biol (2001, 307, 671-681) copyright 2001.

Figures were selected by an automated process.