1OLU

Roles of His291-alpha and His146-beta' in the reductive acylation reaction catalyzed by human branched-chain alpha-ketoacid dehydrogenase

Summary for 1OLU

• Entry DOI: 10.2210/pdb1olu/pdb
• Related: 1DTW 1OLS 1OLX
• Descriptor: 2-OXOISOVALERATE DEHYDROGENASE ALPHA SUBUNIT, 2-OXOISOVALERATE DEHYDROGENASE BETA SUBUNIT, POTASSIUM ION, ... (7 entities in total)
• Functional Keywords: oxidoreductase, ketoacid dehydrogenase, branched-chain, multi-enzyme complex, acylation, oxidative decarboxylation oxidoreductase, maple syrup urine disease, thiamine phosphate
• Biological source: HOMO SAPIENS (HUMAN); More
• Total number of polymer chains: 2
• Total formula weight: 84302.49

Authors

Wynn, R.M.,Machius, M.,Chuang, J.L.,Li, J.,Tomchick, D.R.,Chuang, D.T. (deposition date: 2003-08-15, release date: 2003-08-28, Last modification date: 2023-12-13)

Primary citation

Wynn, R.M.,Machius, M.,Chuang, J.L.,Li, J.,Tomchick, D.R.,Chuang, D.T.
Roles of His291-Alpha and His146-Beta' in the Reductive Acylation Reaction Catalyzed by Human Branched-Chain Alpha-Ketoacid Dehydrogenase: Refined Phosphorylation Loop Structure in the Active Site.
J.Biol.Chem., 278:43402-, 2003

PubMed Abstract: We report here that alterations of either His291-alpha or His146-beta' in the active site of human branched-chain alpha-ketoacid dehydrogenase (E1b) impede both the decarboxylation and the reductive acylation reactions catalyzed by E1b as well as the binding of cofactor thiamin diphosphate (ThDP). In a refined human E1b active-site structure, His291-alpha, which aligns with His407 in Escherichia coli pyruvate dehydrogenase and His263 in yeast transketolase, is on a largely ordered phosphorylation loop. The imidazole ring of His291-alpha in E1b coordinates to the terminal phosphate oxygen atoms of bound ThDP. The N3 atom of wild-type His146-beta', which can be protonated, binds a water molecule and points toward the aminopyrimidine ring of ThDP. Remarkably, the H291A-alpha mutation results in a complete order-to-disorder transition of the loop region, which precludes the binding of the substrate lipoyl-bearing domain to E1b. The H146A-beta' mutation, on the other hand, does not alter the loop structure, but nullifies the reductive acylation activity of E1b. Our results suggest that: 1) His291-alpha plays a structural rather than a catalytic role in the binding of cofactor ThDP and the lipoyl-bearing domain to E1b, and 2) His146-beta' is an essential catalytic residue, probably functioning as a proton donor in the reductive acylation of lipoamide on the lipoyl-bearing domain.

PubMed: 12902323
DOI: 10.1074/JBC.M306204200

Experimental method

X-RAY DIFFRACTION (1.9 Å)