Characterization of human GTPBP3, a GTP-binding protein involved in mitochondrial tRNA modification.
Human GTPBP3 is an evolutionarily conserved, multidomain protein involved in mitochondrial tRNA modification. Characterization of its biochemical properties and the phenotype conferred by GTPBP3 inactivation is crucial to understanding the role of this protein in tRNA maturation and its effects on mitochondrial respiration. We show that the two most abundant GTPBP3 isoforms exhibit moderate affinity for guanine nucleotides like their bacterial homologue, MnmE, although they hydrolyze GTP at a 100-fold lower rate. This suggests that regulation of the GTPase activity, essential for the tRNA modification function of MnmE, is different in GTPBP3. In fact, potassium-induced dimerization of the G domain leads to stimulation of the GTPase activity in MnmE but not in GTPBP3. The GTPBP3 N-terminal domain mediates a potassium-independent dimerization, which appears as an evolutionarily conserved property of the protein family, probably related to the construction of the binding site for the one-carbon-unit donor in the modification reaction. Partial inactivation of GTPBP3 by small interfering RNA reduces oxygen consumption, ATP production, and mitochondrial protein synthesis, while the degradation of these proteins slightly increases. It also results in mitochondria with defective membrane potential and increased superoxide levels. These phenotypic traits suggest that GTPBP3 defects contribute to the pathogenesis of some oxidative phosphorylation diseases.
Villarroya
Magda
M
Laboratorio de Genética Molecular, Centro de Investigación Príncipe Felipe, Avenida Autopista del Saler, 16-3, 46012 Valencia, Spain.
Prado
Silvia
S
Esteve
Juan M
JM
Soriano
Miguel A
MA
Aguado
Carmen
C
Pérez-Martínez
David
D
Martínez-Ferrandis
José I
JI
Yim
Lucía
L
Victor
Victor M
VM
Cebolla
Elvira
E
Montaner
Asunción
A
Knecht
Erwin
E
Armengod
M-Eugenia
ME
D017398:Alternative Splicing; D000818:Animals; D002460:Cell Line; D029968:Escherichia coli Proteins / Q000235:genetics / Q000378:metabolism; D005091:Exons; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D005786:Gene Expression Regulation; D055786:Gene Knockout Techniques; D006150:Guanine Nucleotides / Q000378:metabolism; D006801:Humans; D007438:Introns; D051379:Mice; D008928:Mitochondria / Q000235:genetics*; D015394:Molecular Structure; D010101:Oxygen Consumption; D020033:Protein Isoforms / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D012343:RNA, Transfer / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D029968:Escherichia coli Proteins; D006150:Guanine Nucleotides; D020033:Protein Isoforms; D034741:RNA, Small Interfering; D012343:RNA, Transfer; D020558:GTP Phosphohydrolases; D019204:GTP-Binding Proteins; C498989:GTPBP3 protein, human; C492932:MnmE protein, E coli