NAD: A Renaissance Molecule and its Role in Cell Health

Promega NAD/NADH-Glo system and how to prepare samples for  identification of NAD or NADH.
Promega NAD/NADH-Glo system and how to prepare samples for identification of NAD or NADH.

NAD is a pyridine nucleotide. It provides the oxidation and reduction power for generation of ATP by mitochondria. For many years it was believed that the primary function of NAD/NADH in cells was to harness and transfer energy  from glucose, fatty and amino acids through pathways like glycolysis, beta-oxidation and the citric acid cycle.

Today, however, NAD is recognized as an important cell signaling molecule and substrate. The many regulatory pathways now known to use NAD+ in signaling include multiple aspects of cellular homeostasis, energy metabolism, lifespan regulation, apoptosis, DNA repair and telomere maintenance.

This resurrection of NAD importance is due in no small part to the discovery of NAD-using enzymes, especially the sirtuins. Continue reading “NAD: A Renaissance Molecule and its Role in Cell Health”

Decorating Histones and Their Tails

There are at least two tail stories associated with big scientific discoveries. One is Darwin’s story about the tail loss during human evolution process. The other story is associated with discovery of benzene ring structure. In his creative dream Kekule saw the snake (the linear carbon chain) eating its own tail. Even better representation of benzene structure is the comic image of six monkeys holding each other hands and tails. Nowadays, the most popular scientific story in the field of epigenetics is the story of histones and their tails. This time instead of monkey or snake, an elephant is the animal whose characteristics allegorically represent epigenetics.

elephants_web

We all know how the histone octamer wrapped with DNA represents a nucleosome – the first unit of chromatin formation. Histones, which are basic due to numerous arginines and lysines, easily attract negatively charged DNA and in that way facilitate formation of nucleosome. The nature of two materials is important but not sufficient for such complex biological function like efficient packaging of DNA and regulation of gene expression. For that reason both the DNA and histones are decorated by numerous chemical groups.

Post-translational modifications (PTMs) of histones and histone variants themselves can cause alternation of net charge, changes histone dynamics and interaction with other chromatin proteins. The extreme complexity of interactions that can be achieved by histone modifications inspired Jenuwein and Allis to launch an idea of “histone or epigenetic code”. Core histones consist of a N- terminal tail, the globular portion and a C terminus. PMTs were discovered first on the N-terminal tail of core histones. However, the logical question was: Are only the tails decorated or are there more?” Continue reading “Decorating Histones and Their Tails”

A Scalable and Sensitive Assay for HDAC Activity

Dysfunction of histone deacetylases (HDACs) is associated with many diseases including cancers, asthma and allergies, inflammatory diseases and disorders affecting the central nervous system. Because of their involvement in such a wide range of pathologies, HDACs have become a target for drug discovery. Traditional HDAC activity assays are either isotopic or fluorescent assays using artificial substrates that are prone to artifacts or fluorescence interference. There is a need for a functional assay that is sensitive, accurate and amenable to drug-screening activities.

A recent paper by Halley et al. in the Journal of Biomolecular Screening describes the evaluation of a bioluminogenic HDAC assay, the HDAC-Glo™ I/II and SIRT-Glo™ Assays. Continue reading “A Scalable and Sensitive Assay for HDAC Activity”