1. 8 Protein Interactions with Biomaterials CHAPTER
8.1 Introduction: Thermodynamics of Protein Adsorption
Bacterial surface + Protein adsorption --- Body reaction
Protein adsorption properties ---- Biocompatibility 결정
Composition of proteins and their binding effects on the material surfaces
Gibbs Free Energy and Protein Adsorption
Protein (P) to an empty site on a surface (*): P + * = P*
Gads = Hads – T Sads

2. individual reaction before and after the protein adsorption
for the protein: Gprot = Hprot – T Sprot
for the solvent near the adsorption surface: Gsol = Hsol – T Ssol
for the biomaterial surface: Gsurf = Hsurf – T Ssurf
therefore, Gads = Gprot + Gsol + Gsurf
Systems Properties Governing Protein Adsorption
surface hydrophobicity and charge distribution
factors influencing protein adsorption:
1) dehydration of the surface and protein
2) redistribution of charged groups
3) structural rearrangement of the protein
Interactions between domains
on the biomaterial surface & the protein

4. dehydration for protein adsorption
hydrophobic interaction ---- water disorder ---- entropy 증가
(2) charge interaction
surface --- protein repulsion --- counter ions --- energy expenditure
(3) structural rearrangement of the protein
less stable protein --- preferable adsorption (conformational rearrangement)
8.2. Protein Structure
Amino Acid Chemistry

6. Primary Structure
DNA --- RNA --- Protein
codon usage
missense mutation [HbA & HbS]
Secondary Structure
a-helix
b-sheets [parallel vs. antiparallel]

7. Tertiary Structure
TIM-barrel fold (triosephosphate isomerase)
Noncovalent interactions
ionic interactions / hydrogen bonding /
hydrophobic interaction / van der Waals
Locations of hydrophobic and hydrophilic residues
Quaternary Structure
subunits --- regulatory (allosteric) enzymes
8.3. Protein Transport and Adsorption Kinetics
[prior to the adsorption]
Transport to the Surface
four types: diffusion / thermal convection / flow / coupled transport

8. parabolic velocity profile
ex) blood in the vessel
when r = R, V = 0
---- no flow at the surface
convective and diffusional contributions to overall protein transport
protein transport through a solution + protein adsorption to the surface
with boundary condition
rate of protein adsorption at the surface
= rate of transport of protein to the surface

9. Adsorption Kinetics
biomaterial at the wall --- protein adsorption via diffusion alone
high initial adsorption rate and plateau value
protein rearrangement on the surface
protein monolayer
protein rearrangement
to improve packing

10. 8.4. Reversibility of Protein Adsorption
Reversible & Irreversible Binding
1) short period of reversible binding
initial contact of protein --- conformation change
[protein unfolding and spreading on the material]
2) irreversible (permanent) binding
no desorption [simultaneous desorption]
3) final protein conformation --- activity
Desorption and Exchange
no desorption, but protein exchange
[dynamic nature of protein attachment]
protein displacement
final protein composition on the surface
--- conc. of protein & surface affinity
(size, charge, hydrophobicity,
structural stability, unfolding rate)

11. diffusion-dependent adsorption
--- eventual replacement with greater
surface affinity
[Vroman effect for protein exchange]
8.5. Techniques: Assays for Protein Type and Amount
High-Performance Liquid Chromatography (HPLC):
Affinity Chromatography
Basic principles
adsorption chromatography
normal-phase: polar stationary phase + nonpolar mobile phase
reversed-phase: nonpolar stationary phase + polar mobile phase

12. (2) Instrumentation
detector: UV, refractive index, fluorescence
(3) Information provided
area: quantitative analysis with calibration curve
fractionation (non-destructive monitoring)
Colorimetric Assays
Basic principles and Instrumentation
marker chemical + protein --- chromophore 형성
UV/VIS spectrometry detection (Beer-Lambert’s Law)
product from substrate by enzyme
absorbance monitoring for assessing enzyme activity

13. Fluorescent Assays
Basic principles
fluorophore + protein --- fluorescence
(radiationless decay)
(2) Instrumentation
(3) Information provided
protein quantitation
sensitivity
intrinsic fluorescence (trp, tyr, phe)
Enzyme-linked Immunosorbent Assay (ELISA)
sandwich ELISA
surface - primary Ab – protein – secondary Ab – enzyme
(alkaline phosphatase, horseradish peroxidase, p-nitrophenol phosphatase)

14. Western Blotting
Basic principles and procedures
SDS-PAGE ---- protein transfer to nitrocellulose paper