Two Missense Point Mutations in Different Alleles in the 3-Hydroxy-3-methylglutaryl Coenzyme A Lyase Gene Produce 3-Hydroxy-3-methylglutaric Aciduria in a French Patient

Abstract An alternative target for antifungal drugs is 3-hydroxy-3-methyl glutaryl coenzyme A reductase (HMGR), a key enzyme in the ergosterol biosynthesis pathway. The aim of this study was to obtain, purify, characterize, and overexpress five point mutations in highly conserved regions of the catalytic domain of Candida glabrata HMGR (HMGRCg) to explore the function of key amino acid residues. Glutamic acid (Glu) was substituted by glutamine in the E680Q mutant (at the dimerization site), Glu by glutamine in E711Q (at the substrate binding site), aspartic acid by alanine in D805A and methionine by arginine in M807R (the latter two at the cofactor binding site). A double mutation, E680Q-M807R, was also made. The in vitro enzymatic activity decreased significantly in all recombinant (versus wild-type) HMGRCg, and the in silico binding energy for simvastatin, alpha-asarone and the substrate HMG-CoA was also lower for the mutants. The lowest enzymatic activity and binding energy was displayed by E711Q, suggesting that Glu711 (in the substrate binding site) is an important residue for enzymatic activity. The double mutant HMGRCg E680Q-M807R exhibited the second lowest enzymatic activity. The current findings provide insights into the role of residues in the catalytic site of HMGRCg.

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Characterization of Fenoxaprop-P-Ethyl–Resistant Junglerice (Echinochloa colona) from Mississippi

Weed Science ◽

10.1614/ws-d-16-00024.1 ◽

2016 ◽

Vol 64 (4) ◽

pp. 588-595 ◽

Cited By ~ 6

Author(s):

Alice A. Wright ◽

Vijay K. Nandula ◽

Logan Grier ◽

Kurt C. Showmaker ◽

Jason A. Bond ◽

...

Keyword(s):

Coenzyme A ◽

Point Mutations ◽

Target Site ◽

Acetyl Coenzyme A ◽

Acetyl Coenzyme ◽

Resistant Population ◽

Sunflower County ◽

Acetyl Coenzyme A Carboxylase ◽

P450 Inhibitors

A population of junglerice from Sunflower County, MS, exhibited resistance to fenoxaprop-P-ethyl. An 11-fold difference in ED50 (the effective dose needed to reduce growth by 50%) values was observed when comparing the resistant population (249 g ae ha–1) with susceptible plants (20 g ae ha–1) collected from a different field. The resistant population was controlled by clethodim and sethoxydim at the field rate. Sequencing of the acetyl coenzyme A carboxylase, which encodes the enzyme targeted by fenoxaprop-P-ethyl, did not reveal the presence of any known resistance-conferring point mutations. An enzyme assay confirmed that the acetyl coenzyme A carboxylase in the resistant population is herbicide sensitive. Further investigations with two cytochrome P450 inhibitors, malathion and piperonyl butoxide, and a glutathione-S-transferase inhibitor, 4-chloro-7-nitrobenzofurazan, did not indicate involvement of any metabolic enzymes inhibited by these compounds. The absence of a known target-site point mutation and the sensitivity of the ACCase enzyme to herbicide show that fenoxaprop-P-ethyl resistance in this population is due to a non–target-site mechanism or mechanisms.

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Point mutations in Candida glabrata 3-hydroxy-3-methylglutaryl-coenzyme A reductase (CgHMGR) decrease enzymatic activity and substrate/inhibitor affinity

Scientific Reports ◽

10.1038/s41598-021-00356-w ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Dulce Andrade-Pavón ◽

Vanessa Fernández-Muñoz ◽

Wendy González-Ibarra ◽

César Hernández-Rodríguez ◽

J. Antonio Ibarra ◽

...

Keyword(s):

Binding Energy ◽

Enzymatic Activity ◽

Binding Site ◽

Candida Glabrata ◽

Coenzyme A ◽

Substrate Binding ◽

Point Mutations ◽

Substrate Binding Site ◽

Substrate Inhibitor ◽

Coenzyme A Reductase

Abstract3-Hydroxy-3-methylglutaryl-coenzyme A reductase (HMGR) is a crucial enzyme in the ergosterol biosynthesis pathway. The aim of this study was to obtain, purify, characterize, and overexpress five point mutations in highly conserved regions of the catalytic domain of Candida glabrata HMGR (CgHMGR) to explore the function of key amino acid residues in enzymatic activity. Glutamic acid (Glu) was substituted by glutamine in the E680Q mutant (at the dimerization site), Glu by glutamine in E711Q (at the substrate binding site), aspartic acid by alanine in D805A, and methionine by arginine in M807R (the latter two at the cofactor binding site). A double mutation, E680Q-M807R, was included. Regarding recombinant and wild-type CgHMGR, in vitro enzymatic activity was significantly lower for the former, as was the in silico binding energy of simvastatin, alpha-asarone and the HMG-CoA substrate. E711Q displayed the lowest enzymatic activity and binding energy, suggesting the importance of Glu711 (in the substrate binding site). The double mutant CgHMGR E680Q-M807R exhibited the second lowest enzymatic activity. Based on the values of the kinetic parameters KM and Vmax, the mutated amino acids appear to participate in binding. The current findings provide insights into the role of residues in the catalytic site of CgHMGR.

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Detection of Ki-ras gene point mutations in the bile precipitate of patients with various cause of billiary obstruction

Gastroenterology ◽

10.1016/s0016-5085(01)82854-7 ◽

2001 ◽

Vol 120 (5) ◽

pp. A574-A574

Author(s):

C CHEN ◽

S SHIESH ◽

H TSAO ◽

X LIN

Keyword(s):

Point Mutations ◽

Ras Gene

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Fiberoptic DNA sensor array capable of detecting point mutations

Biosensors and Bioelectronics ◽

10.1016/s0956-5663(97)84360-6 ◽

1998 ◽

Vol 13 (2) ◽

pp. iv-v ◽

Cited By ~ 1

Author(s):

B Healey

Keyword(s):

Sensor Array ◽

Point Mutations ◽

Dna Sensor

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A Novel Mutation Glyl672→Arg in Type 2A and a Homozygous Mutation in Type 2B von Willebrand Disease

Thrombosis and Haemostasis ◽

10.1055/s-0038-1650564 ◽

1996 ◽

Vol 76 (02) ◽

pp. 253-257 ◽

Cited By ~ 14

Author(s):

Takeshi Hagiwara ◽

Hiroshi Inaba ◽

Shinichi Yoshida ◽

Keiko Nagaizumi ◽

Morio Arai ◽

...

Keyword(s):

Missense Mutation ◽

Novel Mutation ◽

Point Mutations ◽

Japanese Patients ◽

Von Willebrand Disease ◽

Homozygous Mutation ◽

Missense Mutations ◽

Reaction Products ◽

Type 3 ◽

Von Willebrand

SummaryGenetic materials from 16 unrelated Japanese patients with von Willebrand disease (vWD) were analyzed for mutations. Exon 28 of the von Willebrand factor (vWF) gene, where point mutations have been found most frequent, was screened by various restriction-enzyme analyses. Six patients were observed to have abnormal restriction patterns. By sequence analyses of the polymerase chain-reaction products, we identified a homozygous R1308C missense mutation in a patient with type 2B vWD; R1597W, R1597Q, G1609R and G1672R missense mutations in five patients with type 2A; and a G1659ter nonsense mutation in a patient with type 3 vWD. The G1672R was a novel missense mutation of the carboxyl-terminal end of the A2 domain. In addition, we detected an A/C polymorphism at nucleotide 4915 with HaeIII. There was no particular linkage disequilibrium of the A/C polymorphism, either with the G/A polymorphism at nucleotide 4391 detected with Hphl or with the C/T at 4891 detected with BstEll.

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