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Catalysts 2016, 6(9), 129; doi:10.3390/catal6090129

Controlling Active Site Loop Dynamics in the (β/α)8 Barrel Enzyme Indole-3-Glycerol Phosphate Synthase

Department of Chemistry, The Pennsylvania State University, University Park, PA 16802, USA
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Academic Editor: Keith Hohn
Received: 9 June 2016 / Revised: 5 August 2016 / Accepted: 22 August 2016 / Published: 26 August 2016
(This article belongs to the Special Issue Enzyme Catalysis)
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Abstract

The β1α1 loop in the tryptophan biosynthetic enzyme indole-3-glycerol phosphate synthase (IGPS) is important for substrate binding, product release and chemical catalysis. IGPS catalyzes the ring closure of the substrate 1-(o-carboxyphenylamine)-1-dexoyribulose 5-phosphate to form indole-3-glycerol phosphate, involving distinct decarboxylation and dehydration steps. The ring closure step is rate-determining in the thermophilic Sulfolobus sulfataricus enzyme (ssIGPS) at high temperatures. The β1α1 loop is especially important in the dehydration step as it houses the general acid Lys53. We propose that loop dynamics are governed by competing interactions on the N- and C-terminal sides of the loop. We had previously shown that disrupting interactions with the N-terminal side of the loop through the N90A substitution decreases catalytic efficiency, slows down the dehydration step and quenches loop dynamics on the picosecond to millisecond timescales. Here, we show that disrupting interactions on the C-terminal side of the loop through the R64A/D65A substitutions likewise decreases catalytic efficiency, slows down the dehydration step and quenches loop dynamics. Interestingly, the triple substitution R64A/D65A/N90A leads to new μs–ms timescale loop dynamics and makes the ring-closure step rate-determining once again. These results are consistent with a model in which the β1α1 loop is maintained in a structurally dynamic state by these competing interactions, which is important for the dehydration step of catalysis. Competing interactions in other enzymes may likewise keep their loops and other structural elements appropriately mobile. View Full-Text
Keywords: enzyme dynamics; enzyme engineering; TIM barrel; indole; tryptophan biosynthesis enzyme dynamics; enzyme engineering; TIM barrel; indole; tryptophan biosynthesis
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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MDPI and ACS Style

O’Rourke, K.F.; Jelowicki, A.M.; Boehr, D.D. Controlling Active Site Loop Dynamics in the (β/α)8 Barrel Enzyme Indole-3-Glycerol Phosphate Synthase. Catalysts 2016, 6, 129.

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