Goldberg 2021 Biochem J

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Goldberg EJ, Buddo KA, McLaughlin KL, Fernandez RF, Pereyra AS, Psaltis CE, Lin CT, Hagen JT, Boykov IN, Nguyen TK, Gowdy KM, Ellis JM, Neufer PD, McClung JM, Fisher-Wellman KH (2021) Tissue-specific characterization of mitochondrial branched-chain keto acid oxidation using a multiplexed assay platform. Biochem J 476:1521-37.

» PMID: 31092703 Open Access

Goldberg Emma J, Buddo Katherine A, McLaughlin Kelsey L, Fernandez Regina F, Pereyra Andrea S, Psaltis Christine E, Lin Chien-Te, Hagen James T, Boykov Ilya N, Nguyen Tiffany K, Gowdy Kymberly M, Ellis Jessica M, Neufer P Darrell, McClung Joseph M, Fisher-Wellman Kelsey H (2021) Biochem J

Abstract: Alterations to branched-chain keto acid (BCKA) oxidation have been implicated in a wide variety of human diseases, ranging from diabetes to cancer. Although global shifts in BCKA metabolism-evident by gene transcription, metabolite profiling, and in vivo flux analyses have been documented across various pathological conditions, the underlying biochemical mechanism(s) within the mitochondrion remain largely unknown. In vitro experiments using isolated mitochondria represent a powerful biochemical tool for elucidating the role of the mitochondrion in driving disease. Such analyses have routinely been utilized across disciplines to shed valuable insight into mitochondrial-linked pathologies. That said, few studies have attempted to model in vitro BCKA oxidation in isolated organelles. The impetus for the present study stemmed from the knowledge that complete oxidation of each of the three BCKAs involves a reaction dependent upon bicarbonate and ATP, both of which are not typically included in respiration experiments. Based on this, it was hypothesized that the inclusion of exogenous bicarbonate and stimulation of respiration using physiological shifts in ATP-free energy, rather than excess ADP, would allow for maximal BCKA-supported respiratory flux in isolated mitochondria. This hypothesis was confirmed in mitochondria from several mouse tissues, including heart, liver and skeletal muscle. What follows is a thorough characterization and validation of a novel biochemical tool for investigating BCKA metabolism in isolated mitochondria. Keywords: Buffer C, Amino acid metabolism, Bioenergetics, Electron transport chain, Mitochondria Bioblast editor: Plangger M, Reiswig R O2k-Network Lab: US NC Greenville Neufer PD


Labels: MiParea: Respiration, Pharmacology;toxicology 


Organism: Mouse  Tissue;cell: Heart, Skeletal muscle, Liver  Preparation: Isolated mitochondria 

Regulation: PCr;Cr  Coupling state: LEAK, OXPHOS, ET  Pathway: F, N, S, NS, ROX  HRR: Oxygraph-2k 

2021-05