Protein Structure Change on Adherence to Ultrafiltration Membranes: An Examination by Electron Paramagnetic Resonance Spectroscopy

✍ Scribed by S.F. Oppenheim; J.O. Rich; G.R. Buettner; V.G.J. Rodgers

• Publisher: Elsevier Science
• Year: 1996
• Tongue: English
• Weight: 134 KB
• Volume: 183
• Category: Article
• ISSN: 0021-9797
• DOI: 10.1006/jcis.1996.0543

✦ Synopsis

also of significant importance. This is primarily because the Electron paramagnetic resonance spectroscopy was used to exadsorbed protein can potentially reduce both sieving and amine changes in the distance between two spin-labels on hen egg permeate flux. However, protein-polymeric interactions lysozyme (HEL, 45 A ˚1 30 A ˚1 30 A ˚, 14,600 Da) when interacting during ultrafiltration may also be different from those prewith membranes during ultrafiltration (UF). Membranes with viously studied. This is primarily because the polymeric nominal molecular weight cutoff (MWCO) values ranging from membranes are porous, resulting in protein interactions un-10,000 to 300,000 Da were used. Both cellulosic (hydrophilic) and der constricted conditions. In addition, the proteinaceous polysulfone (hydrophobic) membranes were studied. The techfluid is driven through the membrane pores by relatively nique used is based on spin-spin interaction of protein-bound large pressure differences, which may promote protein denaspin-labels that cause dipolar broadening of the EPR spectra. This line broadening is related to spin-label distance and was subse-turation.

quently used to infer protein conformational changes. The results

In this work, we investigate protein-membrane interacdemonstrate that the distance between the two labels on HEL, tion as a result of ultrafiltration. Specifically, we will use while in aqueous solution, was approximately 14.4 A ˚. Using the spin-spin interaction of doubly spin-labeled hen egg lysoknown crystal structure of hen egg lysozyme, molecular computer zyme (HEL) to discern tertiary conformational changes. modeling of the doubly labeled protein suggests that the most This method does not require protein desorption from the probable spin-spin distance in solution is 13.9 A ˚. This distance is membrane material. in excellent agreement with our experimentally observed result. Protein-membrane interaction reduced this distance approxi-SPIN-SPIN INTERACTION AND DISTANCE mately by 5 A ˚, irrespective of the membrane morphology. It is speculated that the lack of variation seen in all the membranes, MEASUREMENTS under our test conditions, may be due to limitations associated with spin-label locations.