Flow-induced conformational changes and phase behavior of aqueous poly-L-lysine solutions

Abstract

Poly‐L‐lysine exists as an α‐helix at high pH and a random coil at neutral pH. When the α‐helix is heated above 27°C, the macromolecule undergoes a conformational transition to a β‐sheet. In this study, the stability of the secondary structure of poly‐L‐lysine in solutions subjected to shear flow, at temperatures below the α‐helix to β‐sheet transition temperature, were examined using Raman spectroscopy and CD. Solutions initially in the α‐helical state showed time‐dependent increases in viscosity with shearing, rising as much as an order of magnitude. Visual observation and turbidity measurements showed the formation of a gel‐like phase under flow. Laser Raman measurements demonstrated the presence of small amounts of β‐sheet structure evidenced by the amide I band at 1666 cm−1. CD measurements indicated that solutions of predominantly α‐helical conformation at 20°C transformed into 85% α‐helix and 15% β‐sheet after being sheared for 20 min. However, on continued shearing the content of β‐sheet conformation decreased. The observed phenomena were explained in terms of a “zipping‐up” molecular model based on flow enhanced hydrophobic interactions similar to that observed in gel‐forming flexible polymers. © 1998 John Wiley & Sons, Inc. Biopoly 45: 239–246, 1998

External cavity-quantum cascade laser infrared spectroscopy for secondary structure analysis of proteins at low concentrations

“…In a first step, upon increasing the temperature, intramolecular hydrogen bonds stabilizing the helix configuration break and PLL reversibly adopts this random structure. Further, increasing thermal motion facilitates hydrophobic interactions between lysine side groups, stimulating β-sheet formation, which is irreversible 32 47 . The higher intensity of hydrophobic interactions at elevated temperatures is the basis for the increased propensity for β-sheet formation under these conditions 47 48 .…”

“…Further, increasing thermal motion facilitates hydrophobic interactions between lysine side groups, stimulating β-sheet formation, which is irreversible 32 47 . The higher intensity of hydrophobic interactions at elevated temperatures is the basis for the increased propensity for β-sheet formation under these conditions 47 48 . The positive enthalpy necessary for breaking the hydrogen bonds and association of the lysyl residues forming the β-structure is provided by the heating process 37 .…”

The effects of shear flow on protein structure and function

“…The observed conformational change and subsequent aggregation of the sheared samples was thought to involve flow-enhanced hydrophobic interactions between individual poly-L-lysine molecules in solution. 87 Further studies, via real-time circular birefringence measurements, showed a reversible, shear-induced, helix-to-coil transition of poly-L-lysine in simple shear flow. 24 The conformational transition was observed at a critical strain rate of 300 s 21 , and the change was attributed to a shear-induced breakage of intramolecular hydrogen bonds in a-helical poly-L-lysine.…”

“…87 A combination of Raman spectroscopy (collected in situ) and circular dichroism were employed for data acquisition in a Couette flow cell at 150 s 21 . In addition, a flow induced-gelation and an increase in the b-sheet content of the samples was observed in solution concentrations [0.3 g/dL.…”

Bovine serum albumin unfolds in Couette flow

“…However, a number of experimental and theoretical reports in the literature are yet to reconcile differences between measurement and prediction. For example, flow-induced unfolding of insulin, 21 lysozyme, 15,22 von Willebrand factor, [23][24][25][26][27][28][29][30][31] lamda bacteriophage DNA, 32,33 and several molecular weights of poly-L-lysine 14,34,35 have been reported for a variety of well characterised flow geometries. Shear induced conformational changes in protein solutions have been measured at shear rates <10 3 s À1 (shear strain z10 5 ) while the deformation of DNA coils occurred at even lower shear rates, <4 s À1 (shear strain z10).…”