Dataset Information

Structural and kinetic insights into flavin-containing monooxygenase and calponin-homology domains in human MICAL3.

ABSTRACT: MICAL is an oxidoreductase that participates in cytoskeleton reorganization via actin disassembly in the presence of NADPH. Although three MICALs (MICAL1, MICAL2 and MICAL3) have been identified in mammals, only the structure of mouse MICAL1 has been reported. Here, the first crystal structure of human MICAL3, which contains the flavin-containing monooxygenase (FMO) and calponin-homology (CH) domains, is reported. MICAL3 has an FAD/NADP-binding Rossmann-fold domain for mono-oxygenase activity like MICAL1. The FMO and CH domains of both MICAL3 and MICAL1 are highly similar in structure, but superimposition of the two structures shows a different relative position of the CH domain in the asymmetric unit. Based on kinetic analyses, the catalytic efficiency of MICAL3 dramatically increased on adding F-actin only when the CH domain was available. However, this did not occur when two residues, Glu213 and Arg530, were mutated in the FMO and CH domains, respectively. Overall, MICAL3 is structurally highly similar to MICAL1, which suggests that they may adopt the same catalytic mechanism, but the difference in the relative position of the CH domain produces a difference in F-actin substrate specificity.

SUBMITTER: Kim J

PROVIDER: S-EPMC6949599 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Structural and kinetic insights into flavin-containing monooxygenase and calponin-homology domains in human MICAL3.

Kim Junsoo J Lee Haemin H Roh Yeon Jin YJ Kim Han-Ul HU Shin Donghyuk D Kim Sorah S Son Jonghyeon J Lee Aro A Kim Minseo M Park Junga J Hwang Seong Yun SY Kim Kyunghwan K Lee Yong Kwon YK Jung Hyun Suk HS Hwang Kwang Yeon KY Lee Byung Cheon BC

IUCrJ 20200101 Pt 1

MICAL is an oxidoreductase that participates in cytoskeleton reorganization via actin disassembly in the presence of NADPH. Although three MICALs (MICAL1, MICAL2 and MICAL3) have been identified in mammals, only the structure of mouse MICAL1 has been reported. Here, the first crystal structure of human MICAL3, which contains the flavin-containing monooxygenase (FMO) and calponin-homology (CH) domains, is reported. MICAL3 has an FAD/NADP-binding Rossmann-fold domain for mono-oxygenase activity li ...[more]

PMID: 31949908

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Project description:Busulphan (Bu) is an alkylating agent used in the conditioning regimen prior to hematopoietic stem cell transplantation (HSCT). Bu is extensively metabolized in the liver via conjugations with glutathione to form the intermediate metabolite (sulfonium ion) which subsequently is degraded to tetrahydrothiophene (THT). THT was reported to be oxidized forming THT-1-oxide that is further oxidized to sulfolane and finally 3-hydroxysulfolane. However, the underlying mechanisms for the formation of these metabolites remain poorly understood. In the present study, we performed in vitro and in vivo investigations to elucidate the involvement of flavin-containing monooxygenase-3 (FMO3) and cytochrome P450 enzymes (CYPs) in Bu metabolic pathway. Rapid clearance of THT was observed when incubated with human liver microsomes. Furthermore, among different recombinant microsomal enzymes, the highest intrinsic clearance for THT was obtained via FMO3 followed by several CYPs including 2B6, 2C8, 2C9, 2C19, 2E1 and 3A4. In Bu- or THT-treated mice, inhibition of FMO3 by phenylthiourea significantly suppressed the clearance of both Bu and THT. Moreover, the simultaneous administration of a high dose of THT (200μmol/kg) to Bu-treated mice reduced the clearance of Bu. Consistently, in patients undergoing HSCT, repeated administration of Bu resulted in a significant up-regulation of FMO3 and glutathione-S-transfrase -1 (GSTA1) genes. Finally, in a Bu-treated patient, additional treatment with voriconazole (an antimycotic drug known as an FMO3-substrate) significantly altered the Bu clearance. In conclusion, we demonstrate for the first time that FMO3 along with CYPs contribute a major part in busulphan metabolic pathway and certainly can affect its kinetics. The present results have high clinical impact. Furthermore, these findings might be important for reducing the treatment-related toxicity of Bu, through avoiding interaction with other concomitant used drugs during conditioning and hence improving the clinical outcomes of HSCT.

Dataset Information

Structural and kinetic insights into flavin-containing monooxygenase and calponin-homology domains in human MICAL3.

Publications

Structural and kinetic insights into flavin-containing monooxygenase and calponin-homology domains in human MICAL3.

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