Interaction of nicotinamide adenine dinucleotide and its analogs with glyceraldehyde 3-phosphate dehydrogenase

D, Eby; Me, Kirtley

doi:10.1021/bi00790a004

Cited by 32 publications

(1 citation statement)

References 29 publications

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“…As a cofactor, it binds in the active site and should be a good reporter of protein dynamics in the binding pocket of enzymes. Chromophores substituted at the 3-position of the nicotinamide ring of NAD + are likely to preserve their biological function and bind to the active site in a way similar to that for native NAD + . − Probes that have strong transition moments and transition frequencies that are well separated from other absorption bands are ideal for spectroscopic studies. We have recently shown that an azido-derivatized analog of NAD + , azido-NAD + , has the potential to be a general probe to investigate the active site dynamics of NAD-dependent enzymes .…”

Section: Introductionmentioning

confidence: 99%

3-Picolyl Azide Adenine Dinucleotide as a Probe of Femtosecond to Picosecond Enzyme Dynamics

Dutta

Rock

et al. 2011

J. Phys. Chem. B

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Functionally relevant femtosecond to picosecond dynamics in enzyme active sites can be difficult to measure because of a lack of spectroscopic probes that can be located in the active site without altering the behavior of the enzyme. We have developed a new NAD+ analog 3-Picolyl Azide Adenine Dinucleotide (PAAD+), which has the potential to be a general spectroscopic probe for NAD-dependent enzymes. This analog is stable and binds in the active site of a typical NAD-dependent enzyme formate dehydrogenase (FDH) with similar characteristics to natural NAD+. It has an isolated infrared transition with high molar absorptivity that makes it suitable for observing enzyme dynamics using 2D IR spectroscopy. 2D IR experiments show that in aqueous solution, the analog undergoes complete spectral diffusion within hundreds of femtoseconds consistent with the water hydrogen bonding dynamics that would be expected. When bound to FDH in a binary complex, it shows picosecond fluctuations and a large static offset, consistent with previous studies of the binary complexes of this enzyme. These results show that PAAD+ is an excellent probe of local dynamics and that it should be a general tool for probing the dynamics of a wide range of NAD-dependent enzymes.

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