Substrate-analogue complex structure of Mycobacterium tuberculosis decaprenyl diphosphate synthase

Abstract

Decaprenyl diphosphate synthase from Mycobacterium tuberculosis (MtDPPS, also known as Rv2361c) catalyzes the consecutive elongation of ω,E,Z-farnesyl diphosphate (EZ-FPP) by seven isoprene units by forming new cis double bonds. The protein folds into a butterfly-like homodimer like most other cis-type prenyltransferases. The starting allylic substrate EZ-FPP is bound to the S1 site and the homoallylic substrate to be incorporated, isopentenyl diphosphate, is bound to the S2 site. Here, a 1.55 Å resolution structure of MtDPPS in complex with the substrate analogues geranyl S-thiodiphosphate (GSPP) and isopentenyl S-thiodiphosphate bound to their respective sites in one subunit clearly shows the active-site configuration and the magnesium-coordinated geometry for catalysis. The ligand-binding mode of GSPP in the other subunit indicates a possible pathway of product translocation from the S2 site to the S1 site, as required for the next step of the reaction. The preferred binding of negatively charged effectors to the S1 site also suggests a promising direction for inhibitor design.

Versatile cis-isoprenyl Diphosphate Synthase Superfamily Members in Catalyzing Carbon–Carbon Bond Formation

“…Therefore, D26 was proposed to assist the transfer of Mg 2+ to the allylic substrate. Another feature that has been proposed to play a role in the catalytic reaction is the C-terminal RXG motif, which is located on a loop that extends across the dimer interface and covers the top of the S1 and S2 sites of the other subunit. ,, The guanidinium side chain of the Arg residue provides polar interactions to the diphosphate groups, which is believed to stabilize the reaction complex. , …”

Structural elucidation of the cis -prenyltransferase NgBR/DHDDS complex reveals insights in regulation of protein glycosylation

“…A homozygous mutation in NgBR at Arg-290 (NgBR R290H mutation), occurs in a conserved C-terminal -RXG-motif that is shared amongst NgBR orthologs as well as in bacterial homodimeric cis-PTases, but is absent in DHDDS and its orthologs, suggesting that the C tail of NgBR participates in cis-PTase function. Biochemical characterization of the R290H mutant revealed this mutation impairs IPP binding and markedly reduces catalytic activity but does not influence its interaction with DHDDS 25,33 raising the possibility that NgBR could directly contribute to substrate binding and catalysis through its -RXG-motif 21,43,44 . Another recent exome sequencing study identified multiple mutations in NgBR that appear to cause Parkinson's disease 45 , although it is uncertain whether these mutations affect cis-PTase activity.…”

Structural‐Functional Correlations between Unique N‐terminal Region and C‐terminal Conserved Motif in Short‐chain cis‐Prenyltransferase from Tomato

“…[17] Another crystal structure of DDPS from Mycobacterium tuberculosis suggested that the Cterminal conserved Arg may participate in the transition step of the elongated prenyl diphosphate from the S2 site to the S1 site. [18] However, none of these studies revealed why mutations in the RXG motif result in a significant loss of the condensation activity in the cPT homodimer. L-cPTs utilize heterodimeric systems, and regulatory cPTL proteins contain the RXG motif.…”

“…The crystal structure of dodecaprenyl diphosphate (C 60 ) synthase (DDPS), an M‐cPT from Thermoplasma fusca , indicated the possibility of a negative cooperative mechanism of the RXG motif for the efflux of the diphosphate‐Mg 2+ complex that was generated by carbocation formation of the allylic primer [17] . Another crystal structure of DDPS from Mycobacterium tuberculosis suggested that the C‐terminal conserved Arg may participate in the transition step of the elongated prenyl diphosphate from the S2 site to the S1 site [18] . However, none of these studies revealed why mutations in the RXG motif result in a significant loss of the condensation activity in the cPT homodimer.…”