SensoLyte ® Green Glutaminyl Cyclase Activity Assay *Fluorimetric*
The SensoLyte« Green Glutaminyl Cyclase Activity Assay Kit provides a convenient, two-step homogeneous procedure for measuring enzyme activity from various sources using a green fluorescence substrate. During the first step, substrate is incubated with Glutaminyl Cyclase or enzyme-containing samples and converted into the pyroglutamate form. Secondly, glutaminyl cyclase developer is added to remove pGlu residue and generate the green fluorophore. It can be detected with excitation at 490 nm and emission at 520 nm. Produced fluorescence is proportional to the enzyme activity. This kit is ideal for high throughput screening (HTS) of glutaminyl cyclase activators and inhibitors. The long wavelength of the green fluorophore is less interfered by autofluorescence of cell components and test compounds. Assay sensitivity is 1.5 ng/mL of active enzyme.
Many biologically active peptides and proteins have pyroglutamate (pGlu) as their first amino acid. This modification protects them from degradation by amino peptidases and in many cases determines functionality of protein or peptide.1-6 However, pyroglutamate modification sometimes may lead to negative side effects. For example, pGlu modified beta-amyloid peptides (pGlu-Aβ) aggregate much faster compared to non-pyroglutamic ones and form toxic oligomers that may damage neurons.7-9 Recent studies linked pGlu-Aβ to the Alzheimerĺs Disease because it was identified as a major component of amyloid plaques.7-9
pGlu can be formed when glutamine or glutamic acid spontaneously convert into pyroglutamate. However, cyclic reaction rate is dramatically increased in the presence of Glutaminyl Cyclase (QC), also known as Glutaminyl-peptide Cyclotransferase (QPCT). Human Glutaminyl Cyclase has been isolated from nerve tissue, and detected in cerebrospinal fluid and saliva.6
Figure 1. SensoLyte« Green Glutaminyl Cyclase Assay Principle.
Figure 2.1-Benzyl-Imidazole inhibition of Glutaminyl Cyclase activity measured with SensoLyte« Green Glutaminyl Cyclase Activity Assay Kit (Cat# 72230).
Figure 3. Glutaminyl Cyclase Activity in Human Body Fluids. Human cerebrospinal fluid (CSF) and saliva were diluted 2.5-fold using assay buffer ±Glutaminyl Cyclase inhibitor and were tested for the enzyme activity using SensoLyte« Green Glutaminyl Cyclase Assay Kit (Cat# 72230).