title: Concentrating membrane proteins using ultrafiltration without concentrating detergents.
abstract: Membrane proteins (MPs) are of rapidly growing interest in the design of pharmaceutical products, novel sensors, and synthetic membranes. Ultrafiltration (UF) using commercially available centrifugal concentrators is typically employed for laboratory-scale concentration of low-yield MPs, but its use is accompanied by a concomitant increase in concentration of detergent micelles. We present a detailed analysis of the hydrodynamic processes that control detergent passage during ultrafiltration of MPs and propose methods to optimize detergent passage during protein concentration in larger-scale membrane processes. Experiments were conducted using nonionic detergents, octyl-β-D glucoside (OG), and decyl-β-D maltoside (DM) with the bacterial water channel protein, Aquaporin Z (AqpZ) and the light driven chloride pump, halorhodopsin (HR), respectively. The observed sieving coefficient (So ), a measure of detergent passage, was evaluated in both stirred cell and centrifugal systems. So for DM and OG increased with increasing filtrate flux and decreasing shear rates in the stirred cell, that is, with increasing concentration polarization (CP). Similar effects were observed during filtration of MP-detergent (MPD) micelles. However, lower transmission was observed in the centrifugal system for both detergent and MPD systems. This is attributed to free convection-induced shear and hence reduced CP along the membrane surface during centrifugal UF. Thus to concentrate MPs without retention of detergent, design of UF systems that promote CP is required. Biotechnol. Bioeng. 2016;113: 2122-2130. © 2016 Wiley Periodicals, Inc.
authors: Department of Chemical Engineering, The Pennsylvania State University, 155 Fenske Laboratory, University Park, Pennsylvania, 16802.
authors: Hasin
authors: H
authors: Feroz
authors: Department of Chemical Engineering, The Pennsylvania State University, 155 Fenske Laboratory, University Park, Pennsylvania, 16802.
authors: Craig
authors: C
authors: Vandervelden
authors: Department of Chemical Engineering, The Pennsylvania State University, 155 Fenske Laboratory, University Park, Pennsylvania, 16802.
authors: Bon
authors: B
authors: Ikwuagwu
authors: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania.
authors: Bryan
authors: B
authors: Ferlez
authors: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania.
authors: Carol S
authors: CS
authors: Baker
authors: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania.
authors: Daniel J
authors: DJ
authors: Lugar
authors: Applied Biomimetic Inc., Cincinnati, Ohio.
authors: Mariusz
authors: M
authors: Grzelakowski
authors: Department of Biochemistry and Molecular Biology, The Pennsylvania State University, University Park, Pennsylvania.|Department of Chemistry, The Pennsylvania State University, University Park, Pennsylvania.
authors: John H
authors: JH
authors: Golbeck
authors: Department of Chemical Engineering, The Pennsylvania State University, 155 Fenske Laboratory, University Park, Pennsylvania, 16802.
authors: Andrew L
authors: AL
authors: Zydney
authors: Department of Chemical Engineering, The Pennsylvania State University, 155 Fenske Laboratory, University Park, Pennsylvania, 16802. manish.kumar@psu.edu.
authors: Manish
authors: M
authors: Kumar
mesh_terms: D002498:Centrifugation / Q000295:instrumentation* / Q000379:methods*; D003902:Detergents / Q000737:chemistry*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D008565:Membrane Proteins / Q000302:isolation & purification*; D014462:Ultrafiltration / Q000295:instrumentation* / Q000379:methods*
chemical_list: D003902:Detergents; D008565:Membrane Proteins
keywords: concentration polarization; membrane proteins; ultrafiltration