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Frontal affinity ch...
Frontal affinity chromatographic analysis of membrane protein reconstitution
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- Lundqvist, Andreas (author)
- Uppsala universitet,Institutionen för naturvetenskaplig biokemi
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- Brekkan, Eggert (author)
- Uppsala universitet,Institutionen för naturvetenskaplig biokemi
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- Lagerquist, Christine (author)
- Uppsala universitet,Institutionen för naturvetenskaplig biokemi
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- Haneskog, Lars (author)
- Uppsala universitet,Institutionen för naturvetenskaplig biokemi
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- Lundahl, Per (author)
- Uppsala universitet,Institutionen för naturvetenskaplig biokemi
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(creator_code:org_t)
- 1997
- 1997
- English.
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In: Materials science & engineering. C, biomimetic materials, sensors and systems. - 0928-4931 .- 1873-0191. ; 4:4, s. 221-226
- Related links:
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https://urn.kb.se/re...
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https://doi.org/10.1...
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Abstract
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- The human red cell glucose transporter Glut1 was solubilized with octaoxyethylene n-dodecyl ether (low critical micelle concentration (CMC)), purified, mixed with egg phospholipids and cholate, and reconstituted by gel filtration on Superdex 75. Free protepliposomes showed relatively high D-glucose transport activity. Frontal affinity chromatographic analysis with the proteoliposomes sterically immobilized in Superdex 200 gel beads revealed that the number of operative cytochalasin B (CB) binding sites increased during the first days of chromatographic runs to become the same as with 1-O-n-octyl β-d-glucopyranoside (high CMC) as solubilizer and Sephadex G-50 as gel filtration medium. The average number of sites per Glut1 monomer was 0.32 ± 0.02. The average Kd for CB was 66 ± 3 nM at 150 mM NaCl, similarly as for Glut1 in membrane vesicles, whereas the affinity of d-glucose for reconstituted Glut1 was lower (Kd = 44 ± 3 mM) than for membranous Glut1 (Kd = 15 ± 5 mM). Two theoretical treatments of affinity chromatographic data gave the same values in agreement with competitive and monovalent interactions.
Publication and Content Type
- ref (subject category)
- art (subject category)
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