1VG3

Crystal Structure Of Octaprenyl Pyrophosphate Synthase From Hyperthermophilic Thermotoga Maritima A76Y/S77F mutant

Summary for 1VG3

• Entry DOI: 10.2210/pdb1vg3/pdb
• Related: 1V4E 1V4H 1V4I 1V4J 1V4K 1VG2 1VG4 1VG6 1VG7
• Descriptor: octoprenyl-diphosphate synthase, SULFATE ION (3 entities in total)
• Functional Keywords: trans-type prenyltransferase, thermophilic, transferase
• Biological source: Thermotoga maritima
• Total number of polymer chains: 1
• Total formula weight: 34148.59

Authors

Guo, R.T.,Kuo, C.J.,Ko, T.P.,Chou, C.C.,Liang, P.H.,Wang, A.H.-J. (deposition date: 2004-04-23, release date: 2004-05-18, Last modification date: 2023-10-25)

Primary citation

Guo, R.T.,Kuo, C.J.,Ko, T.P.,Chou, C.C.,Liang, P.H.,Wang, A.H.-J.
A molecular ruler for chain elongation catalyzed by octaprenyl pyrophosphate synthase and its structure-based engineering to produce unprecedented long chain trans-prenyl products
Biochemistry, 43:7678-7686, 2004

PubMed Abstract: Octaprenyl pyrophosphate synthase (OPPs) catalyzes consecutive condensation reactions of farnesyl pyrophosphate (FPP) with five molecules of isopentenyl pyrophosphate (IPP) to generate C(40) octaprenyl pyrophosphate (OPP) which constitutes the side chain of menaquinone. We have previously reported the X-ray structure of OPPs from Thermotoga maritima, which is composed entirely of alpha-helices joined by connecting loops and is arranged with nine core helices around a large central cavity [Guo, R. T., Kuo, C. J., Ko, T. P., Chou, C. C., Shr, R. L., Liang, P. H., and Wang, A. H.-J. (2004) J. Biol. Chem. 279, 4903-4912]. A76 and S77 are located on top of the active site close to where FPP is bound. A76Y and A76Y/S77F OPPs mutants produce C(20), indicating that the substituted larger residues interfere with the substrate chain elongation. Surprisingly, the A76Y/S77F mutant synthesizes a larger amount of C(20) than the A76Y mutant. In the crystal structure of the A76Y/S77F mutant, F77 is pushed away by Y76, thereby creating more space between those two large amino acids to accommodate the C(20) product. A large F132 residue at the bottom of the tunnel-shaped active site serves as the "floor" and determines the final product chain length. The substitution of F132 with a small Ala, thereby removing the blockade, led to the synthesis of a C(50) product larger than that produced by the wild-type enzyme. On the basis of the structure, we have sequentially mutated the large amino acids, including F132, L128, I123, and D62, to Ala underneath the tunnel. The products of the F132A/L128A/I123A/D62A mutant reach C(95), beyond the largest chain length generated by all known trans-prenyltransferases. Further modifications of the enzyme reaction conditions, including new IPP derivatives, may allow the preparation of high-molecular weight polyprenyl products resembling the rubber molecule.

PubMed: 15196010
DOI: 10.1021/bi036336d

Experimental method

X-RAY DIFFRACTION (2.7 Å)