Harnessing Liquid Crystal Sensors for High-Throughput Real-Time Detection of Structural Changes in Lysozyme during Refolding Processes.

Despite the rapid advances in process analytical technology, the assessment of protein refolding efficiency has largely relied on off-line protein-specific assays and/or chromatographic procedures such as reversed-phase high-performance liquid chromatography and size exclusion chromatography. Due to the inherent time gap pertaining to traditional methods, exploring optimum refolding conditions for many recombinant proteins, often expressed as insoluble inclusion bodies, has proven challenging. The present study describes a novel protein refolding sensor that utilizes liquid crystals (LCs) to discriminate varying protein structures during unfolding and refolding. An LC layer containing 4-cyano-4'-pentylbiphenyl (5CB) intercalated with 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) is used as a sensing platform, and its proof-of-concept performance is demonstrated using lysozyme as a model protein. As proteins unfold or refold, a local charge fluctuation at their surfaces modulates their interaction with zwitterionic phospholipid DOPE. This alters the alignment of DOPE molecules at the aqueous/LC interface, affecting the orientational ordering of bulk LC (i.e., homeotropic to planar for refolding and planar to homeotropic for unfolding). Differential polarized optical microscope images of the LC layer are subsequently generated, whose brightness directly linked to conformational changes of lysozyme molecules is quantified by gray scale analysis. Importantly, our LC-based refolding sensor is compatible with diverse refolding milieus for real-time analysis of lysozyme refolding and thus likely to facilitate the refolding studies of many proteins, especially those lacking a method to determine structure-dependent biological activity.

Fabrication of Liquid Crystal Droplet Patterns for Monitoring Aldehyde Vapors.

A sensing method based on the pattern of liquid crystal droplets was developed for detecting and monitoring low levels of organic aldehyde vapors. Exposure of the LC droplet pattern covered with glycine solution to aldehyde vapors induced an optical signal transition from a bright fan shape to a dark cross appearance, as observed by polarized light microscopy. Aldehyde and glycine react at the air/solution interface to form a Schiff-base compound, which controls the orientation of the LCs and induces a change in the optical signals of the LC droplet pattern. The results show that the glycine/LC droplet pattern system is particularly sensitive and selective to aldehydes. In the actual environment, the sensor is exposed to the aldehyde and the signal transition is completed within a few minutes (2-7 min). The LC-based method has the advantages of simple construction, easy operation, convenient data reading, and shows excellent prospects for real-time detection of aldehyde vapors.