title: Reengineering of the human pyruvate dehydrogenase complex: from disintegration to highly active agglomerates.
abstract: The pyruvate dehydrogenase complex (PDC) plays a central role in cellular metabolism and regulation. As a metabolite-channeling multi-enzyme complex it acts as a complete nanomachine due to its unique geometry and by coupling a cascade of catalytic reactions using 'swinging arms'. Mammalian and specifically human PDC (hPDC) is assembled from multiple copies of E1 and E3 bound to a large E2/E3BP 60-meric core. A less restrictive and smaller catalytic core, which is still active, is highly desired for both fundamental research on channeling mechanisms and also to create a basis for further modification and engineering of new enzyme cascades. Here, we present the first experimental results of the successful disintegration of the E2/E3BP core while retaining its activity. This was achieved by C-terminal α-helixes double truncations (eight residues from E2 and seven residues from E3BP). Disintegration of the hPDC core via double truncations led to the formation of highly active (approximately 70% of wildtype) apparently unordered clusters or agglomerates and inactive non-agglomerated species (hexamer/trimer). After additional deletion of N-terminal 'swinging arms', the aforementioned C-terminal truncations also caused the formation of agglomerates of minimized E2/E3BP complexes. It is likely that these 'swinging arm' regions are not solely responsible for the formation of the large agglomerates.
authors.affiliation: Institute of Bioprocess and Biosystems Engineering, Technische Universität Hamburg, 21073 Hamburg, Germany.
authors.forename: Jin
authors.initials: J
authors.lastname: Guo
authors.affiliation: Institute of Bioprocess and Biosystems Engineering, Technische Universität Hamburg, 21073 Hamburg, Germany.
authors.forename: Samira
authors.initials: S
authors.lastname: Hezaveh
authors.affiliation: Institute of Bioprocess and Biosystems Engineering, Technische Universität Hamburg, 21073 Hamburg, Germany.
authors.forename: Jana
authors.initials: J
authors.lastname: Tatur
authors.affiliation: Institute of Bioprocess and Biosystems Engineering, Technische Universität Hamburg, 21073 Hamburg, Germany.
authors.forename: An-Ping
authors.initials: AP
authors.lastname: Zeng
authors.affiliation: Institute of Bioprocess and Biosystems Engineering, Technische Universität Hamburg, 21073 Hamburg, Germany uwe.jandt@tuhh.de.
authors.forename: Uwe
authors.initials: U
authors.lastname: Jandt
mesh_terms: D000105:Acetyl Coenzyme A / Q000737:chemistry* / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D020134:Catalytic Domain; D003001:Cloning, Molecular; D008058:Dihydrolipoamide Dehydrogenase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051047:Dihydrolipoyllysine-Residue Acetyltransferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D007700:Kinetics; D009154:Mutation; D015202:Protein Engineering; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D030481:Pyruvate Dehydrogenase (Lipoamide) / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011768:Pyruvate Dehydrogenase Complex / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D019289:Pyruvic Acid / Q000737:chemistry* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013329:Structure-Activity Relationship
chemical_list: D011768:Pyruvate Dehydrogenase Complex; D011994:Recombinant Proteins; D000105:Acetyl Coenzyme A; D019289:Pyruvic Acid; D030481:Pyruvate Dehydrogenase (Lipoamide); D008058:Dihydrolipoamide Dehydrogenase; D051047:Dihydrolipoyllysine-Residue Acetyltransferase