Andrographolide inhibits human serum albumin fibril formations through site-specific molecular interactions

Abstract

Protein misfolding and fibrillation are the fundamental traits in degenerative diseases like Alzheimer's, Parkinsonism, and diabetes mellitus. Bioactives such as flavonoids and terpenoids from plant sources are known to express protective effects against an array of diseases including diabetes, Alzheimer's and obesity. Andrographolide (AG), a labdane diterpenoid is prescribed widely in the Indian and Chinese health care systems for classical efficacy against a number of degenerative diseases. This work presents an in depth study on the effects of AG on protein fibrillating pathophysiology. Thioflavin T fluorescence spectroscopy and DLS results indicated concentration dependent inhibition of human serum albumin (HSA) fibrillation. The results were confirmed by electron microscopy studies. HSA fibril formations were markedly reduced in the presence of AG. Fluorescence studies and UV-Vis experiments confirmed further that AG molecularly interacts with HSA at site. In silico molecular docking studies revealed hydrogen bonding and hydrophobic interactions with HSA in the native state. Thus AG interacts with HSA, stabilizes the native protein structure and inhibits fibrillation. The results demonstrated that the compound possesses anti-amyloidogenic properties and can be promising against some human degenerative diseases.

Fig. 1. Fibril growth of HSA in absence and presence of different ratios of AG as measured by DLS at 25 °C.

Fig. 2. ThT growth curve of HSA in absence and presence of different ratios of AG.

Fig. 3. (A) Absorption spectra of AG (a), HSA in absence (b) and presence of different concentration of AG (c–f) (B) fluorescence spectra of HSA in presence of different concentration of AG (a–e).

Fig. 4. (A) Chemical structure of andrographolide (B) surface view of the crystal structure of HSA (PDB ID: 2BXP) bound to docked andrographolide showing H-bond (yellow dotted lines) interactions with binding site residue.

Fig. 5. TEM images of 2 μM HSA solution incubated alone (A) and with AG at ratios of (B) 1 : 0.5, (C) 1 : 1, (D) 1 : 2. Scale: 0.2 μm.

Fig. 6. (A) Far UV CD spectra and (B) histograms of % β-sheet content in HSA solution in absence and presence of AG.