PMID- 26575208
OWN - NLM
STAT- MEDLINE
DCOM- 20160411
LR  - 20211203
IS  - 1520-4995 (Electronic)
IS  - 0006-2960 (Print)
IS  - 0006-2960 (Linking)
VI  - 54
IP  - 49
DP  - 2015 Dec 15
TI  - Mechanistic Investigation of cPMP Synthase in Molybdenum Cofactor Biosynthesis 
      Using an Uncleavable Substrate Analogue.
PG  - 7229-36
LID - 10.1021/acs.biochem.5b00857 [doi]
AB  - Molybdenum cofactor (Moco) is essential for all kingdoms of life, plays central 
      roles in various biological processes, and must be biosynthesized de novo. During 
      its biosynthesis, the characteristic pyranopterin ring is constructed by a 
      complex rearrangement of guanosine 5'-triphosphate (GTP) into cyclic pyranopterin 
      monophosphate (cPMP) through the action of two enzymes, MoaA and MoaC. Recent 
      studies revealed that MoaC catalyzes the majority of the transformation and 
      produces cPMP from a unique cyclic nucleotide, 3',8-cyclo-7,8-dihydro-GTP 
      (3',8-cH2GTP). However, the mechanism by which MoaC catalyzes this complex 
      rearrangement is largely unexplored. Here, we report the mechanistic 
      characterization of MoaC using an uncleavable substrate analogue, 
      3',8-cH2GMP[CH2]PP, as a probe to investigate the timing of cyclic phosphate 
      formation. Using partially active MoaC variants, 3',8-cH2GMP[CH2]PP was found to 
      be accepted by MoaC as a substrate and was converted to an analogue of the 
      previously described MoaC reaction intermediate, suggesting that the early stage 
      of catalysis proceeds without cyclic phosphate formation. In contrast, when it 
      was incubated with wt-MoaC, 3',8-cH2GMP[CH2]PP caused mechanism-based inhibition. 
      Detailed characterization of the inhibited MoaC suggested that 3',8-cH2GMP[CH2]PP 
      is mainly converted to a molecule (compound Y) with an acid-labile 
      triaminopyrimidinone base without an established pyranopterin structure. MS 
      analysis of MoaC treated with 3',8-cH2GMP[CH2]PP provided strong evidence that 
      compound Y forms a tight complex with MoaC likely through a covalent linkage. 
      These observations are consistent with a mechanism in which cyclic phosphate ring 
      formation proceeds in concert with the pterin ring formation. This mechanism 
      would provide a thermodynamic driving force to complete the formation of the 
      unique tetracyclic structure of cPMP.
FAU - Hover, Bradley M
AU  - Hover BM
AD  - Department of Biochemistry, Duke University Medical Center , Durham, North 
      Carolina 27710, United States.
FAU - Lilla, Edward A
AU  - Lilla EA
AD  - Department of Biochemistry, Duke University Medical Center , Durham, North 
      Carolina 27710, United States.
FAU - Yokoyama, Kenichi
AU  - Yokoyama K
AD  - Department of Biochemistry, Duke University Medical Center , Durham, North 
      Carolina 27710, United States.
LA  - eng
GR  - R01 GM112838/GM/NIGMS NIH HHS/United States
PT  - Journal Article
PT  - Research Support, N.I.H., Extramural
PT  - Research Support, Non-U.S. Gov't
DEP - 20151201
PL  - United States
TA  - Biochemistry
JT  - Biochemistry
JID - 0370623
RN  - 0 (Coenzymes)
RN  - 0 (Escherichia coli Proteins)
RN  - 0 (Metalloproteins)
RN  - 0 (MoaC protein, E coli)
RN  - 0 (Molybdenum Cofactors)
RN  - 0 (Organophosphorus Compounds)
RN  - 0 (Pteridines)
RN  - 0 (Pterins)
RN  - 0 (nulibry)
RN  - ATN6EG42UQ (molybdenum cofactor)
RN  - EC 3.- (Hydrolases)
RN  - EC 3.3.1.2 (S-adenosylmethionine enzyme MoaA, Staphylococcus aureus)
SB  - IM
MH  - Coenzymes/biosynthesis/*chemistry
MH  - Escherichia coli/*enzymology
MH  - Escherichia coli Proteins/*chemistry/metabolism
MH  - Hydrolases/chemistry/metabolism
MH  - Mass Spectrometry
MH  - Metalloproteins/biosynthesis/*chemistry
MH  - Molybdenum Cofactors
MH  - Organophosphorus Compounds/*chemistry/metabolism
MH  - Pteridines/*chemistry
MH  - Pterins/*chemistry/metabolism
MH  - Staphylococcus aureus/enzymology
PMC - PMC4847533
MID - NIHMS776048
EDAT- 2015/11/18 06:00
MHDA- 2016/04/12 06:00
PMCR- 2016/04/27
CRDT- 2015/11/18 06:00
PHST- 2015/11/18 06:00 [entrez]
PHST- 2015/11/18 06:00 [pubmed]
PHST- 2016/04/12 06:00 [medline]
PHST- 2016/04/27 00:00 [pmc-release]
AID - 10.1021/acs.biochem.5b00857 [doi]
PST - ppublish
SO  - Biochemistry. 2015 Dec 15;54(49):7229-36. doi: 10.1021/acs.biochem.5b00857. Epub 
      2015 Dec 1.