Substrate Channeling via a Transient Protein-Protein Complex: The Case of D-Glyceraldehyde-3-Phosphate Dehydrogenase and L-Lactate Dehydrogenase

Zeljko Svedruzic, Ivica Odorcic, Christopher Chang, Drazenka Svedruzic

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10 Scopus Citations

Abstract

Substrate channeling studies have frequently failed to provide conclusive results due to poor understanding of this subtle phenomenon. We analyzed the mechanism of NADH-channeling from D-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) to L-lactate Dehydrogenase (LDH) using enzymes from different cells. Enzyme kinetics studies showed that LDH activity with free NADH and GAPDH-NADH complex always take place in parallel. The channeling is observed only in assays that mimic cytosolic conditions where free NADH concentration is negligible and the GAPDH-NADH complex is dominant. Molecular dynamics and protein-protein interaction studies showed that LDH and GAPDH can form a leaky channeling complex only at the limiting NADH concentrations. Surface calculations showed that positive electric field between the NAD(H) binding sites on LDH and GAPDH tetramers can merge in the LDH-GAPDH complex. NAD(H)-channeling within the LDH-GAPDH complex can be an extension of NAD(H)-channeling within each tetramer. In the case of a transient LDH-(GAPDH-NADH) complex, the relative contribution from the channeled and the diffusive paths depends on the overlap between the off-rates for the LDH-(GAPDH-NADH) complex and the GAPDH-NADH complex. Molecular evolution or metabolic engineering protocols can exploit substrate channeling for metabolic flux control by fine-tuning substrate-binding affinity for the key enzymes in the competing reaction paths.

Original languageAmerican English
Article numberArticle No. 10404
Number of pages12
JournalScientific Reports
Volume10
Issue number1
DOIs
StatePublished - 1 Dec 2020

Bibliographical note

Publisher Copyright:
© 2020, The Author(s).

NREL Publication Number

  • NREL/JA-2700-73920

Keywords

  • glyceraldehyde-3-phosphate dehydrogenase
  • lactate dehydrogenase
  • substrate channeling

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