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lecture6 - 3.051J/20.340J 1 Lecture 6 Protein-Surface...

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1 3.051 J / 20 .340 J Lecture 6: Protein-Surface Interactions ( Part II) The Langmuir model is applicable to numerous reversible adsorption processes, but fails to capture many aspects of protein adsorption. 1. Competitive Adsorption ¾ many different globular proteins in vivo ¾ surface distribution depends on [P i ]’s & time The Vroman effect : Displacement (over time) of initially adsorbed protein by a second protein. S.L. Cooper, J. Biomater. Sci. 3 , 1991: 27-47.) Γ FGN Γ FN Γ VN Γ ( 2 ) 0 300 200 100 FGN, FN, VN adsorption on a polyether urethane from plasma (from D.J. Fabrizius-Homan & ng/cm 60 120 time (min) Protein Plasma conc. (mg/ml) MW (Daltons) Human serum albumin 42 68,500 Immunoglobulins 28 145,000 (IgG) Fibrinogen 3.0 340,000 Fibronectin 0.3 240,000 Vitronectin 0.2 60,000 Plasma – fluid component of blood with anticoagulant added Serum – fluid component of blood with coagulants removed
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2 3.051 J / 20 .340 J Hypothesis: At t~0 : uniform [P i ]’s everywhere protein with highest concentration dominates initial adsorption At t>0: local depletion of adsorbed species near surface– exchange with faster diffusing species ensues At t>>0: gradual exchange with higher affinity species 2. Irreversible Adsorption ¾ occurs in vivo & in vitro: proteins often do not desorb after prolonged exposure to protein solutions ¾ complicates the competitive adsorption picture % FGN remaining 100 PDMS Glass Surfaces exposed to plasma after of FGN adsorption (from S.M. Slack and T.A. 0 FGN adsorp. time (min) 60 Horbett, J. Colloid & Intfc Sci . 133 , 1989: 148.)
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3 3.051 J / 20 .340 J Physiological implications: a) hydrophobic surfaces cause more denaturing b) denatured proteins may ultimately desorb (by replacement) non-native solution behavior Models that attempt to account for 1 & 2: S.M. Slack and T.A. Horbett, J. Colloid & Intfc Sci . 133, 1989 p. 148 I. Lundstroem and H. Elwing, J. Colloid & Intfc Sci . 136, 1990 p. 68 C.F. Lu, A. Nadarajah, and K.K. Chittur, J. Coll. & Intfc Sci . 168, 1994 p. 152 3. Restructuring ¾ Protein layers reaching monolayer saturation can reorganize (e.g., crystallize) on surface, creating a stepped isotherm Γ time
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4 3.051 J / 20 .340 J 4. Multilayer Formation ¾ Proteins can adsorb atop protein monolayers or sublayers, creating complicated adsorption profiles Γ time
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5 3.051 J / 20 .340 J Measurement of Adsorbed Proteins 1. Techniques for Quantifying Adsorbed Amount a) Labeling Methods : tag protein for quantification, use known standards for calibration i) Radioisotopic labeling ¾ proteins labeled with radioactive isotopes that react with specific a.a. residues e.g., tyrosine labeling with 125 I ; 131 I; 32 P OH - CH 2 125 I
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