Two-photon excited-state and conformation dynamics of NADH binding with dehydrogenases

Yuexin Liu, Florly S. Ariola, Hye Ryong Kim, Qianru Yu, Walvick Ronn, Heikal Ahmed

Research output: Chapter in Book/Report/Conference proceedingChapter

2 Scopus citations


Reduced β-nicotinamide adenine dinucleotide (NADH) is the principal electron donor in glycolytic and oxidative energy metabolism in living cells. Distinctive differences between the excited-state dynamics of free NADH in solution and its intrinsic counterpart in living cells or brain slices are indicated. This chapter investigates the two-photon fluorescence lifetime and rotational anisotropy of NADH, under equilibrium binding conditions with malate dehydrogenase (MDH) and lactate dehydrogenase (LDH), in solution to reconcile these differences. The results indicate an enhancement of the fluorescence lifetime of NADH by a factor of ∼ 2 upon MDH enzyme binding, which is consistent with literature values using other biochemical techniques. Furthermore, equilibrated NADH-MDH reveals a complex rotational anisotropy at enzyme concentrations around the K. d value. Analogous measurements on NADH binding with LDH reveal similar trends, albeit different kinetic parameters. The results are compared with lipoamide dehydrogenase where flavin adenine dinucleotide is covalently bound to the enzyme backbone.

Original languageEnglish (US)
Title of host publicationFemtochemistry VII
Number of pages6
ISBN (Print)9780444528216
StatePublished - 2006
Externally publishedYes

Bibliographical note

Funding Information:
This work was supported by the National Science Foundation (MRSEC/PSU, COT/Lehigh, and MRI/PSU) and the Huck Institutes of the Life Sciences (PSU). HK was supported by BBSI (NIH and NSF undergraduate program; Bioengineering, PSU). The authors also express their gratitude to Coherent Lasers, Inc. for their loan of a pulse picker (MIRA9200, Coherent).


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