Bovine and human insulin adsorption at lipid monolayers: a comparison

Abstract

Insulin is a widely used peptide in protein research and it is utilized as a model peptide to understand the mechanics of fibril formation, which is believed to be the cause of diseases such as Alzheimer and Creutzfeld-Jakob syndrome. Insulin has been used as a model system due to its biomedical relevance, small size and relatively simple tertiary structure. The adsorption of insulin on a variety of surfaces has become the focus of numerous studies lately. These works have helped in elucidating the consequence of surface/protein hydrophilic/hydrophobic interaction in terms of protein refolding and aggregation. Unfortunately, such model surfaces differ significantly from physiological surfaces. Here we spectroscopically investigate the adsorption of insulin at lipid monolayers, to further our understanding of the interaction of insulin with biological surfaces. In particular we study the effect of minor mutations of insulin's primary amino acid sequence on its interaction with 1,2-Dipalmitoyl-sn-glycero-3-phosphoglycerol (DPPG) model lipid layers. We probe the structure of bovine and human insulin at the lipid/water interface using sum frequency generation spectroscopy (SFG). The SFG experiments are complemented with XPS analysis of Langmuir-Schaefer deposited lipid/insulin films. We find that bovine and human insulin, even though very similar in sequence, show a substantially different behavior when interacting with lipid films.

The air-water interface stabilizes α-helical conformations of the insulin B-chain

“…This is likely a general phenomenon but extension of this to other proteins and to other interfaces will be necessary to determine specific driving forces and how this is affected by protein structure and environment. For instance simulation of insulin on lipid monolayers may be used to resolve the differences in insulin behaviour at more complex interfaces compared to the bare AWI 34,79 . This may then be used to give further insight into the mechanisms of protein aggregation, such as the formation of protein fibrils, which may be used to guide and control the formation of protein aggregates.…”

“…The formation of α-helical structures at the air-water interface is consistent with SFG measurements of insulin at the bare air-water interface 34 . In comparison at lipid monolayers the SFG signal for human insulin is reduced 79 , suggesting either an increased tendency for the formation of SFG inactive dimers or that the protein adopts more disordered conformations. This suggests that the hydrophobicity of the interface plays a key role in determining protein interfacial conformation and aggregation.…”

Otoferlin C2F Domain-Induced Changes in Membrane Structure Observed by Sum Frequency Generation

“…To identify how the protein influences the structure of the lipid layer it is interacting with, SFG spectra were collected at a lipid monolayer surface both before and after the C2F domain was allowed to interact for 2.5 h. Lipid monolayers provide a simple biomimetic approach to study the structure and interaction of proteins at the lipid membrane interface (14,15,24,32,(47)(48)(49)(50)(51)(52)(53). Again, as in previous studies, isotopically labeled acyl chain versions of the lipids isolate the vibrations of just the lipid monolayers acyl chains for that of the proteins vibrational CH groups (14,15).…”

“…Protein SFG spectra are heavily influenced by interference between signals from different secondary-structure elements and protein side chains, based on their relative orientation and energy. SFG has previously been used to study lipid monolayers at the air/water interface (15,32,(49)(50)(51)(52)57,58), model lipid bilayers (59)(60)(61)(62), and small proteins and peptides interaction with each type of model membrane (14)(15)(16)53,59,(61)(62)(63)(64)(65)(66)(67)(68)(69)(70). For small proteins and peptides, the direct analysis of SFG amide-I spectra by peak fitting vibrational spectra relating to the C¼O of the amide backbone can provide information about the orientation and structure (67,71,72).…”

<p>Size-Dependent Interactions of Lipid-Coated Gold Nanoparticles: Developing a Better Mechanistic Understanding Through Model Cell Membranes and in vivo Toxicity</p>

“…One approach to a systematic study is through the use of well-studied biomimetic lipid monolayers that have been employed to assess the interactions of biomolecules or NPs with a lipid monolayer. [32][33][34][35][36][37][38][39][40][41] Structural changes in the biomimetic lipid monolayer are monitored by sum-frequency generation (SFG) vibrational spectroscopy, a second-order nonlinear optical technique. SFG is capable of detecting the adsorption and orientation of biomolecules at sub-micromolar concentrations.…”