Structural targeting of a familial variant of neuroserpin (Ser52Arg) by epigallocatechin gallate binding that reduces polymerization and enhances the tissue plasminogen activator inhibition.

Neuroserpin (NS) inactivation of tissue plasminogen activator (tPA) in brain reduces neurotoxicity. However several familial point mutations in its gene are linked with a neurodegenerative disease termed Familial Encephalopathy with Neuroserpin Inclusion Bodies (FENIB) that results in epilepsy/dementia. Variable expression level of the NS has been linked to several other neurological pathologies like Alzheimer's, glaucoma and ischemic stroke. A familial variant of NS that leads to mortality, Ser52Arg (S52R) was made using site directed mutagenesis. A circular dichroism study indicated close correspondence in the secondary structures of S52R and the recombinant NS. However, fluorometric analysis showed a conformational deformation and a shift towards more hydrophobic environment. Consequently, S52R showed an enhanced ability to form polymers based on the native PAGE and Thioflavin T binding studies. Epigallocatechin gallate (EGCG), a polyphenol with neuroprotective properties, is shown to have a high affinity for both, the NS and the S52R in a comprehensive screening of the natural compounds. Interestingly, S52R showed reduction in the polymer formation and significant retention of its tPA inhibition activity on incubation with ECGC (100 μM). A site specific labelling of S52 using Alexa fluor 488C5 maleimide dye indicated that this region undergoes burial on addition of the tPA. A guanidium hydrochloride based denaturation study showed that EGCG increases the conformational stability of a folding intermediate. EGCG binds to the native NS, a folding intermediate and the natural variant of NS and retards the polymer formation with significant retention of tPA inhibition activity with implications in reducing the pathological symptoms.