ライフサイエンスコーパス: AdoCbl

1 AdoCbl (5'-deoxyadenosylcobalamin) is synthesized from c

2 AdoCbl produced by R. sphaeroides was identified and qua

3 Coenzyme B(12) (AdoCbl; 5'-deoxy-5'-adenosylcobalamin), the quintessenti

4 nts of cob(II)alamin generated with [5'-2H2]-AdoCbl in D2O in comparison with AdoCbl in H2O reveal tw

5 th [5'-2H2]-AdoCbl in H2O, and with [5'-2H2]-AdoCbl in D2O.

6 has now been measured in D2O, with [5'-2H2]-AdoCbl in H2O, and with [5'-2H2]-AdoCbl in D2O.

7 talyzes the exchange of tritium from [5'-3H]-AdoCbl with solvent.

8 dual functions; i.e., it tailors the active AdoCbl form of the cofactor and then transfers it direct

9 lonyl-CoA mutase is an 5'-adenosylcobalamin (AdoCbl)-dependent enzyme that catalyzes the rearrangemen

10 e catalytic cycles of all adenosylcobalamin (AdoCbl)-dependent enzymes, as in each case catalysis is

11 dical propagation from an adenosylcobalamin (AdoCbl) to a pyridoxal 5'-phosphate (PLP) cofactor.

12 thine 4,5-aminomutase, an adenosylcobalamin (AdoCbl)- and pyridoxal L-phosphate (PLP)-dependent enzym

13 lmalonyl-CoA mutase is an adenosylcobalamin (AdoCbl)-dependent enzyme that catalyzes the rearrangemen

14 yl-CoA mutase (MUT) is an adenosylcobalamin (AdoCbl)-requiring mitochondrial matrix enzyme that catal

15 4,5-aminomutase (OAM), an adenosylcobalamin (AdoCbl; coenzyme B(12))-dependent isomerase, employs a l

16 coenzymes for 5,6-LAM are adenosylcobalamin (AdoCbl) and pyridoxal 5'-phosphate (PLP).

17 es and uses coenzyme B12, adenosylcobalamin (AdoCbl), as a cofactor.

18 ethanolamine catalyzed by adenosylcobalamin (AdoCbl)-dependent ethanolamine ammonia-lyase (EAL).

19 lus leichmannii catalyzes adenosylcobalamin (AdoCbl)-dependent nucleotide reduction, as well as excha

20 ed to bring the cofactors adenosylcobalamin (AdoCbl) and pyridoxal-5'-phosphate (PLP) and the substra

21 erivatization to 5'-deoxy-adenosylcobalamin (AdoCbl), the required MUT cofactor.

22 metry and apparent Kd for adenosylcobalamin (AdoCbl) are dependent upon the relative concentrations o

23 spectrum is observed for adenosylcobalamin (AdoCbl) in water and ethylene glycol.

24 ydrase and the functional adenosylcobalamin (AdoCbl) analogue 5'-deoxy-3',4'-anhydroadenosylcobalamin

25 cteria, the late steps in adenosylcobalamin (AdoCbl) biosynthesis are collectively known as the nucle

26 he lower ligand, which in adenosylcobalamin (AdoCbl) is 5,6-dimethylbenzimidazole, and in adenosylpse

27 air (RP) stabilization in adenosylcobalamin (AdoCbl)-dependent enzymes and (ii) the manifestation of

28 etalloproteins, including adenosylcobalamin (AdoCbl)-dependent methylmalonyl-CoA mutase and hydrogena

29 to biological activity of adenosylcobalamin (AdoCbl) and methylcobalamin (MeCbl) is the Co-C bond cle

30 of the nucleotide loop of adenosylcobalamin (AdoCbl) and other cobamides.

31 the carbon-cobalt bond of adenosylcobalamin (AdoCbl) at a rate approximately 10(11)-fold faster than

32 zation of the kinetics of adenosylcobalamin (AdoCbl) binding by stopped-flow fluorescence spectroscop

33 te in the biosynthesis of adenosylcobalamin (AdoCbl) in many prokaryotes.

34 acilitate the delivery of adenosylcobalamin (AdoCbl) to AdoCbl-dependent methylmalonyl-CoA mutase, an

35 ilitating the delivery of adenosylcobalamin (AdoCbl) to methylmalonyl-CoA mutase (MCM), the only AdoC

36 e de novo biosynthesis of adenosylcobalamin (AdoCbl), catalyzing the formation of the essential Co-C

37 e belongs to the class of adenosylcobalamin (AdoCbl)-dependent carbon skeleton isomerases and catalyz

38 d for the reactivation of adenosylcobalamin (AdoCbl)-dependent diol dehydratase.

39 ed in the biosynthesis of adenosylcobalamin (AdoCbl).

40 catalyze the formation of adenosylcobalamin (AdoCbl, coenzyme B(12)) from cobalamin and ATP.

41 ide (Cbi), a precursor of adenosylcobalamin (AdoCbl, coenzyme B(12)).

42 d for the biosynthesis of adenosylcobalamin (AdoCbl; coenzyme B(12) ).

43 ulsed-laser photolysis of adenosylcobalamin (AdoCbl; coenzyme B(12)) in AdoCbl-dependent ethanolamine

44 hat delivers the product, adenosylcobalamin (AdoCbl or coenzyme B(12)), to methylmalonyl-CoA mutase (

45 of ethanolamine requires adenosylcobalamin (AdoCbl) as a cofactor, and, intriguingly, we also identi

46 lass II RNR that requires adenosylcobalamin (AdoCbl) as a cofactor.

47 bl) is an analogue of the adenosylcobalamin (AdoCbl) coenzyme.

48 enzyme that utilizes the adenosylcobalamin (AdoCbl) cofactor to catalyze the rearrangement of methyl

49 The adenosylcobalamin (AdoCbl)-dependent enzyme ethanolamine ammonia-lyase (EAL

50 reaction catalyzed by the adenosylcobalamin (AdoCbl)-dependent enzyme, methylmalonyl-CoA mutase, has

51 nosyl radical pair in the adenosylcobalamin (AdoCbl)-dependent ethanolamine ammonia-lyase (EAL) from

52 y, studied herein are the adenosylcobalamin (AdoCbl, also known as coenzyme B(12))-dependent diol deh

53 i, a 76 kDa monomer using adenosylcobalamin (AdoCbl) as a cofactor, catalyzes the conversion of nucle

54 llus leichmannii utilizes adenosylcobalamin (AdoCbl) as a cofactor and, in addition to nucleotide red

55 (12) into coenzyme B(12) (adenosylcobalamin, AdoCbl) is catalyzed by ATP:cob(I)alamin adenosyltransfe

56 sm by which coenzyme B12 (adenosylcobalamin, AdoCbl)-dependent enzymes promote homolytic cleavage of

57 ,2-PD) in a coenzyme B12 (adenosylcobalamin, AdoCbl)-dependent fashion.

58 rate, and product in the adenosylcobalamin- (AdoCbl) dependent reaction of ethanolamine ammonia-lyase

59 As is the case for all AdoCbl-dependent isomerases, the catalytic cycle of EAL

60 es of Cbls is Ado-cobinamide (AdoCbi(+)), an AdoCbl derivative that lacks the tethered base 5,6-dimet

61 ding motif, the first to be discovered in an AdoCbl-dependent enzyme.

62 ase and acted as an in vitro inhibitor of an AdoCbl-dependent diol dehydratase.

63 l that the "unique" Abs spectra of MeCbl and AdoCbl, which differ considerably from the "typical" Abs

64 phosphate, and isotopically labeled RTPR and AdoCbl in conjunction with EPR spectroscopy has allowed

65 re the molecular basis of G-protein-assisted AdoCbl-dependent mutase maturation, explaining how GTP b

66 resence of methylmalonyl-CoA mutase and ATP, AdoCbl is transferred from ATR to the acceptor protein i

67 r B12 biochemistry and renders coenzyme B12 (AdoCbl) so intriguingly suitable for enzymatic radical r

68 ever, EAL and other Class II isomerases bind AdoCbl in the so-called "base-on" conformation and must

69 This protein, GlmES, binds AdoCbl stoichiometrically and neither the affinity for A

70 ared to be from the reaction of NO with both AdoCbl intermediates (Cbl(II) and .CH(2)-Ado) generated

71 Structural models of free and EAL-bound AdoCbl were constructed using molecular dynamics and qua

72 for Co-C bond cleavage of free and EAL-bound AdoCbl, we identified key cofactor/enzyme interactions t

73 rate of Co-C bond homolysis of enzyme-bound AdoCbl is increased by 12 orders of magnitude.

74 es rotation and a ~14 A translation to bring AdoCbl-initially positioned ~25 A away-into the active-s

75 studies reveal novel strategies employed by AdoCbl-dependent enzymes in the control of radical catal

76 sites in ATR are simultaneously occupied by AdoCbl.

77 r forms methyl-Cbl (MeCbl) and adenosyl-Cbl (AdoCbl) is required for the function of two crucial enzy

78 r of the enzyme, 5'-deoxyadenosyl-cobalamin (AdoCbl), on substrate binding.

79 e regulated by a single adenosyl cobalamine (AdoCbl)-responsive riboswitch.

80 reconstituted with both the native cofactor AdoCbl and its derivative methylcobalamin (MeCbl).

81 actor of 4 compared with the native cofactor AdoCbl.

82 y weakened affinity for the native cofactor, AdoCbl.

83 In this solvent-exposed conformation, AdoCbl undergoes facile transfer to MCM.

84 The light-responsive cobalt-corrin AdoCbl performs this nonenzymatic role by facilitating t

85 ylmalonic acidemia, resulting from defective AdoCbl binding.

86 , which also serves as an escort, delivering AdoCbl to methylmalonyl-CoA mutase (MCM).

87 uccinyl-CoA using 5'-deoxyadenosylcobalamin (AdoCbl) as a metallocofactor.

88 te catalysis, the 5'-deoxyadenosylcobalamin (AdoCbl) cofactor's Co-C bond is cleaved homolytically to

89 he mutase for the 5'-deoxyadenosylcobalamin (AdoCbl) cofactor, increasing it 2-fold from 404 +/- 71 t

90 lamin (MeCbl) and 5'-deoxyadenosylcobalamin (AdoCbl) have long fascinated chemists because of their c

91 min, synthesizing 5'-deoxyadenosylcobalamin (AdoCbl) or coenzyme B(12).

92 coenzyme B(12) or 5'-deoxyadenosylcobalamin (AdoCbl), which serves as a cofactor for a number of isom

93 IcmF is a 5'-deoxyadenosylcobalamin (AdoCbl)-dependent enzyme that catalyzes the carbon skele

94 5'-Deoxyadenosylcobalamin (AdoCbl)-dependent isomerases catalyze carbon skeleton re

95 of coenzyme B12 (5'-deoxyadenosylcobalamin, AdoCbl), in which the configuration of the N-glycosidic

96 pulsed-laser photolysis of AdoCbl in the EAL-AdoCbl-substrate ternary complex, and time-resolved prob

97 We show that AdoRhbl closely emulates AdoCbl in its uptake by bacterial cells and structural f

98 by significantly distorting the equilibrium AdoCbl structure.

99 ATR is also a chaperone that escorts AdoCbl, transferring it to methylmalonyl-CoA mutase, whi

100 ucture different from previously established AdoCbl riboswitches.

101 Compared with wild type, the affinity for AdoCbl is unchanged, but for the conversion of L-glutama

102 ichiometrically and neither the affinity for AdoCbl nor the turnover number depends upon protein conc

103 SR reduced cob(II)alamin to cob(I)alamin for AdoCbl synthesis (in conjunction with the prior finding

104 idues of switch III as a wedge, allowing for AdoCbl insertion or damaged Cbl removal.

105 quadrant of the molecule, as is the case for AdoCbl.

106 t to keep up with even the lowest demand for AdoCbl.

107 The mutations increased the K(m) for AdoCbl by 40- to 900-fold, while V(max) values varied fr

108 g that EutT employs a distinct mechanism for AdoCbl formation.

109 alue (245 microM compared to 0.54 microM for AdoCbl).

110 The binding mode for AdoCbl is notably different from that of cob(II)alamin,

111 excited-state spectrum is also observed for AdoCbl bound to glutamate mutase.

112 lizes a conformation of MCM that is open for AdoCbl insertion, and GTP hydrolysis, as signaled by swi

113 as a photostable isostructural surrogate for AdoCbl.

114 8 +/- 0.01, which is 3-fold smaller than for AdoCbl in aqueous solution (0.23 +/- 0.01).

115 ibo)AdoCbl is 160-fold smaller than that for AdoCbl, and only 1/3 as much cob(II)alamin is produced a

116 A comparison of these results with those for AdoCbl in H2O revealed kH/kD of 1.6, 1.7, and 2.7, respe

117 e, only two of its active sites are used for AdoCbl synthesis and where binding of ATP to the vacant

118 that PduX is an L-threonine kinase used for AdoCbl synthesis.

119 , which attacks the 5'-carbon of ATP to form AdoCbl and inorganic triphosphate.

120 pKa of 3.7 for dimethylbenzimidazole in free AdoCbl.

121 the continuous-wave C-Co photolysis of free AdoCbl in 75% glycerol but have not done so in the therm

122 de that removal of the phosphoryl group from AdoCbl-5'-P is the last step in AdoCbl biosynthesis in s

123 terface, thus uncoupling GTP hydrolysis from AdoCbl delivery.

124 For Class I AdoCbl-dependent isomerases, an important contribution t

125 of radical-based catalysis in this Class III AdoCbl-dependent enzyme.

126 In AdoCbl-depleted enzyme, formation of the external aldimi

127 denosylcobalamin (AdoCbl; coenzyme B(12)) in AdoCbl-dependent ethanolamine ammonia-lyase (EAL) from S

128 we considered the substitution of cobalt in AdoCbl with rhodium to generate the rhodium analogue 5'-

129 tion reaction from a primary CH(3)- group in AdoCbl-dependent methylmalonyl-CoA mutase shows the enzy

130 otated as cobY to reflect its involvement in AdoCbl biosynthesis.

131 l group from AdoCbl-5'-P is the last step in AdoCbl biosynthesis in serovar Typhimurium and that the

132 C-C bond angle are quite similar to those in AdoCbl.

133 c chromatin loop with LRRK2, whose inhibitor AdoCbl could be an anti-tuberculosis drug candidate.

134 ls the Rossmann domain, harboring the intact AdoCbl cofactor, is tilted toward the edge of the PLP bi

135 iguingly, we also identify an intercistronic AdoCbl riboswitch that has a predicted structure differe

136 tial for the conversion of AdopseudoCbl into AdoCbl, the cobamide needed for the catabolism of acetat

137 e incomplete corrinoid cobinamide (Cbi) into AdoCbl.

138 o their upper axial ligand, including MeCbl, AdoCbl, aquacobalamin (H(2)OCbl(+)), and vitamin B(12) (

139 In the proposed chemical mechanism, AdoCbl initiates the formation of substrate radicals, an

140 variants have sufficient activity to mediate AdoCbl synthesis in vivo.

141 steric effector dGTP (Km = 17 +/- 3 microM), AdoCbl (Km = 60 +/- 9 microM) and no external reductant.

142 me uses AdopseudoCbl as a substrate, but not AdoCbl.

143 phate (AdoCbi-P), an intermediate in de novo AdoCbl biosynthesis.

144 ekeeping enzyme that is required for de novo AdoCbl synthesis and for salvaging incomplete precursors

145 Salmonella obtains AdoCbl by assimilation of complex precursors, such as vi

146 In the absence of AdoCbl, EutX uses this structure to sequester EutV.

147 AdoRhbl, the non-natural rhodium analogue of AdoCbl, as a photostable isostructural surrogate for Ado

148 We demonstrate that association of AdoCbl to this riboswitch prevents formation of an intri

149 hydrolyzable analogue of GTP, the binding of AdoCbl to the mutase is not detected.

150 It is proposed that the polyhedra consist of AdoCbl-dependent diol dehydratase (and perhaps other pro

151 Spectroscopic data of AdoCbl and Co(II)Cbl show minimal perturbations upon cof

152 previously demonstrated that only 2 equiv of AdoCbl bind per homotrimer of ATR and that binding of AT

153 active site triggers ejection of 1 equiv of AdoCbl from an adjacent site.

154 ATR required for delivery of 1 equivalent of AdoCbl, from 4 to 1.

155 fectively compete with the back formation of AdoCbl.

156 in formed have been studied as a function of AdoCbl concentration and temperature.

157 at L-2-hydroxyglutarate-induced homolysis of AdoCbl occurs very rapidly, with a rate constant approac

158 , as well as exchange of the 5' hydrogens of AdoCbl with solvent.

159 to precisely target biomimetic inhibition of AdoCbl-based photoregulation, with new possibilities for

160 visible stopped-flow kinetic measurements of AdoCbl homolysis obtained with deuterated substrates.

161 oupled to repositioning of the Ado moiety of AdoCbl from the eastern conformation to the northern con

162 (13)C introduced into the ribosyl moiety of AdoCbl.

163 proximately 7 A from its position as part of AdoCbl to a position where it is in contact with C1 of t

164 Photolysis of AdoCbl in EAL at 240 K leads to cob(II)alamin-5'-deoxyad

165 Photolysis of AdoCbl in EAL leads to a quantum yield at 10(-7) s for c

166 studied by using pulsed-laser photolysis of AdoCbl in the EAL-AdoCbl-substrate ternary complex, and

167 pha-ribazole-5'-P, alpha-RP), a precursor of AdoCbl.

168 arable with that observed in the presence of AdoCbl (5.0 +/- 0.6) and indicates that the hydrogen tra

169 ady-state conditions reveals the presence of AdoCbl but no cob(II)alamin.

170 In the presence of AdoCbl, EutV cannot bind to EutX and, instead, causes tr

171 uctures of IcmF, a natural fusion protein of AdoCbl-dependent isobutyryl-CoA mutase and its correspon

172 m the active site, precluding reformation of AdoCbl at the end of the turnover cycle.

173 roism (CD), and magnetic CD (MCD) spectra of AdoCbl are observed upon formation of holoenzyme, even i

174 ferase, which catalyzes the terminal step of AdoCbl synthesis.

175 73) and is required for de novo synthesis of AdoCbl (coenzyme B(12)).

176 dent enzymes or for the de novo synthesis of AdoCbl.

177 ith maximal activity that is 9.7% of that of AdoCbl itself, and a very high Km value (245 microM comp

178 in ring is considerably flatter than that of AdoCbl, with a fold angle of 11.7 degrees.

179 oRbl revealed a structure similar to that of AdoCbl.

180 ofactor ejection, which leads to transfer of AdoCbl from wild-type ATR to MCM.

181 (beta), which forms a part of the "base-off"-AdoCbl binding motif.

182 at synthesizes GkCblS ectopically makes only AdoCbl, even under growth conditions where the synthesis

183 to methylmalonyl-CoA mutase (MCM), the only AdoCbl-dependent enzyme in humans.

184 e much smaller than those measured for other AdoCbl enzymes and model reactions for which hydrogen tu

185 erases tend to be specialized for particular AdoCbl-dependent enzymes or for the de novo synthesis of

186 hosphorylate adenosylcobalamin-5'-phosphate (AdoCbl-5'-P), the product of the condensation of alpha-r

187 gest that the adenosyl group of photoexcited AdoCbl bound to CarH may specifically undergo a concerte

188 ulated the enzymatic reaction by photolyzing AdoCbl, and found that even at low NO concentrations, NO

189 thermolysis of their respective precursors, AdoCbl and 8-MeOAdoCbl.

190 ld, respectively), affinity for the product, AdoCbl, is significantly diminished (400-fold), and the

191 Cloning and expression of the putative AdoCbl-dependent PCM with an alpha2beta2 heterotetrameri

192 rturbs the equilibrium between the reactant (AdoCbl-bound) state and the product (cob(II)alamin/5'-de

193 TR signals that its cofactor cargo is ready (AdoCbl) or not [cob(II)alamin] for transfer to MCM, is n

194 rch, the mechanism by which MMCM and related AdoCbl-dependent enzymes accelerate the rate for homolyt

195 of alleles (class M) that failed to restore AdoCbl biosynthesis during intragenic complementation st

196 the L. innocua cblT and cblS genes restored AdoCbl synthesis from Cbi and alpha-R in a Salmonella en

197 op assembly pathway in cobC strains restored AdoCbl-5'-P synthesis from Cby in a cobC strain.

198 d carbon-cobalt bond cleavage of (alpha-ribo)AdoCbl is 160-fold smaller than that for AdoCbl, and onl

199 d in the Ado ligand is inverted [(alpha-ribo)AdoCbl], has been synthesized and its crystal structure

200 from samples prepared with [U-(13)C-ribosyl]-AdoCbl.

201 tant results from its inability to sequester AdoCbl for direct transfer to MCM.

202 ty; thus, variant proteins failed to support AdoCbl synthesis in vivo.

203 iant enzymes analyzed in this work supported AdoCbl biosynthesis in vivo.

204 that R. sphaeroides strain 2.4.1 synthesizes AdoCbl de novo and that it salvages Cbi using both of th

205 on, and immunoelectron microscopy shows that AdoCbl-dependent diol dehydratase is associated with the

206 ties for the cob(II)alamin substrate and the AdoCbl product and increase K(M(ATP)).

207 um and that the reaction is catalyzed by the AdoCbl-5'-P phosphatase (CobC) enzyme.

208 (alpha-RP) and adenosylcobinamide-GDP by the AdoCbl-5'-P synthase (CobS, EC 2.7.8.26) enzyme.

209 o PLP as an external aldimine and elicit the AdoCbl Co-C bond homolysis and the accumulations of cob(

210 of cofactor loading and offloading from the AdoCbl-dependent IcmF are distinct from those of the bet

211 ture, which is independent of changes in the AdoCbl structure, and specifically the Co-C bond length,

212 ic acid (DAB) induces rapid homolysis of the AdoCbl Co-C bond (781 s(-1), D-ornithine; 513 s(-1), DAB

213 ogether demonstrate unusual stability of the AdoCbl Co-C bond and that radical catalysis is coupled t

214 n, which is critical in the formation of the AdoCbl Co-C bond.

215 termediate required for the formation of the AdoCbl product.

216 otein that comprises the two subunits of the AdoCbl-dependent isobutyryl-CoA mutase flanking a G-prot

217 We report that the AdoCbl-binding riboswitch is part of a small, trans-acti

218 As compared to the AdoCbl data, however, Abs and MCD spectral changes for t

219 In common with the AdoCbl-dependent enzymes catalyzing irreversible heteroa

220 to light triggers the decomposition of this AdoCbl-bound complex by a still elusive photochemical me

221 min to cob(I)alamin that was adenosylated to AdoCbl by ATR.

222 to Co2+ to Co+, followed by adenosylation to AdoCbl.

223 he delivery of adenosylcobalamin (AdoCbl) to AdoCbl-dependent methylmalonyl-CoA mutase, an essential

224 ransferase in turn converts cob(II)alamin to AdoCbl in the presence of ATP and a reductant.

225 th purified ATR can convert cob(II)alamin to AdoCbl in vitro.

226 inactive cobalamins, such as vitamin B12, to AdoCbl.

227 tly the conversion of inactive cobalamins to AdoCbl for 1,2-propanediol degradation.

228 y which this metallochaperone contributes to AdoCbl delivery without directly binding the cofactor.

229 homologues represent previously unidentified AdoCbl-dependent enzymes is discussed.

230 places the pendant base from the Co ion upon AdoCbl binding to these enzymes.

231 site leads to the transfer of the high value AdoCbl product to the acceptor mutase.

232 s accelerated by 12 orders of magnitude when AdoCbl is bound to the protein active site, possibly thr

233 ingly, our previous studies showed that when AdoCbl is bound to the MMCM active site, no enzymatic pe

234 n as the global minimum, as is the case with AdoCbl itself.

235 th [5'-2H2]-AdoCbl in D2O in comparison with AdoCbl in H2O reveal twice as much cob(II)alamin in the

236 bstrate ATP to ATR that is fully loaded with AdoCbl leads to the ejection of 1 equivalent of the cofa

237 iol dehydrase at 0.02% of that observed with AdoCbl.

238 To study the mechanism of NO reaction with AdoCbl, we simulated the enzymatic reaction by photolyzi

239 by the sugar of F(2)CTP and the second with AdoCbl destruction.

240 f growth on similar medium supplemented with AdoCbl.