title: pH-responsive modulation of insulin aggregation and structural transformation of the aggregates.
abstract: Over the past two decades, much information has appeared on electrostatically driven molecular mechanisms of protein self-assembly and formation of aggregates of different morphology, varying from soluble amorphous structures to highly-ordered amyloid-like fibrils. Protein aggregation represents a special tool in biomedicine and biotechnology to produce biological materials for a wide range of applications. This has awakened interest in identification of pH-triggered regulators of transformation of aggregation-prone proteins into structures of higher order. The objective of the present study is to elucidate the effects of low-molecular-weight biogenic agents on aggregation and formation of supramolecular structures of human recombinant insulin, as a model therapeutic protein. Using dynamic light scattering, turbidimetry, circular dichroism, fluorescence spectroscopy, atomic force microscopy, transmission electron microscopy, and nuclear magnetic resonance, we have demonstrated that the amino acid l-arginine (Arg) has the striking potential to influence insulin aggregation propensity. It was shown that modification of the net charge of insulin induced by changes in the pH level of the incubation medium results in dramatic changes in the interaction of the protein with Arg. We have revealed the dual effects of Arg, highly dependent on the pH level of the solution - suppression or acceleration of the aggregation of insulin at pH 7.0 or 8.0, respectively. These effects can be regulated by manipulating the pH of the environment. The results of this study may be of interest for development of appropriate drug formulations and for the more general insight into the functioning of insulin in living systems, as the protein is known to release by exocytosis from pancreatic beta cells in a pH-dependent manner.
authors.affiliation: A.N. Bach Institute of Biochemistry, Russian Academy of Sciences, Leninsky Prospect, 33, 119071 Moscow, Russia.
authors.forename: Ekaterina
authors.initials: E
authors.lastname: Smirnova
authors.affiliation: A.N. Bach Institute of Biochemistry, Russian Academy of Sciences, Leninsky Prospect, 33, 119071 Moscow, Russia.
authors.forename: Irina
authors.initials: I
authors.lastname: Safenkova
authors.affiliation: A.N. Bach Institute of Biochemistry, Russian Academy of Sciences, Leninsky Prospect, 33, 119071 Moscow, Russia.
authors.forename: Vita
authors.initials: V
authors.lastname: Stein-Margolina
authors.affiliation: A.N. Bach Institute of Biochemistry, Russian Academy of Sciences, Leninsky Prospect, 33, 119071 Moscow, Russia.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Shubin
authors.affiliation: Center for Magnetic Tomography & Spectroscopy, Faculty of Fundamental Medicine, M.V. Lomonosov Moscow State University, Vorobievy Gory, GSP-1, Moscow 119991, Russia.
authors.forename: Vladimir
authors.initials: V
authors.lastname: Polshakov
authors.affiliation: A.N. Bach Institute of Biochemistry, Russian Academy of Sciences, Leninsky Prospect, 33, 119071 Moscow, Russia. Electronic address: bella@inbi.ras.ru.
authors.forename: Bella
authors.initials: B
authors.lastname: Gurvits
mesh_terms: D001120:Arginine / Q000737:chemistry* / Q000378:metabolism; D002942:Circular Dichroism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007328:Insulin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy; D018625:Microscopy, Atomic Force; D046529:Microscopy, Electron, Transmission; D009391:Nephelometry and Turbidimetry; D066329:Protein Aggregates; D011994:Recombinant Proteins / Q000737:chemistry* / Q000378:metabolism; D013050:Spectrometry, Fluorescence
chemical_list: D007328:Insulin; D066329:Protein Aggregates; D011994:Recombinant Proteins; D001120:Arginine
keywords: Arginine; Atomic force microscope; Insulin; Protein aggregation; pH-regulation