He Huang

Protein adsorption on poly(N-vinylpyrrolidone)-modified silicon surfaces prepared by surface-initiated atom transfer radical polymerization.

Well-controlled poly(N-vinylpyrrolidone) (PVP)-grafted silicon surfaces were prepared by surface-initiated atom transfer radical polymerization (SI-ATRP) with 1,4-dioxane/water mixtures as solvents and CuCl/5,7,7,12,14,14-hexamethyl-1,4,8,11-tetraazacyclotetradecane (Me6TATD) as a catalyst. The thickness of the PVP layer on the surface increased with reaction time, suggesting that the ATRP grafting of N-vinylpyrrolidone (NVP) from the silicon surfaces was a well-controlled process. The water contact angle and X-ray photoelectron spectroscopy (XPS) were used to characterize the modified surfaces. The protein adsorption property of the PVP-grafted surfaces was evaluated using a radiolabeling method. Compared with unmodified silicon surfaces, a Si-PVP60 surface with a PVP thickness of 15.06 nm reduced the level of adsorption of fibrinogen, human serum albumin (HSA), and lysozyme by 75, 93, and 81%, respectively. Moreover, the level of fibrinogen adsorption decreases gradually with an increase in PVP thickness. However, no significant difference in fibrinogen adsorption was found when the PVP layer was thicker than the critical thickness of 13.45 nm.

Protein adsorption and cell adhesion/detachment behavior on dual-responsive silicon surfaces modified with poly(N-isopropylacrylamide)-block-polystyrene copolymer.

Diblock copolymer grafts covalently attached to surfaces have attracted considerable attention because of their special structure and novel properties. In this work, poly(N-isopropylacrylamide)-block-polystyrene (PNIPAAm-b-PS) brushes were prepared via surface-initiated consecutive atom-transfer radical polymerization on initiator-immobilized silicon. Because of the inherent thermosensitivity of PNIPAAm and the hydrophobicity difference between the two blocks, the modified surfaces were responsive to both temperature and solvent. Moreover, the diblock copolymer brushes exhibited both resistance to nonspecific protein adsorption and unique cell interaction properties. They showed strong protein resistance in both phosphate-buffered saline and blood plasma. In particular, fibrinogen adsorption from plasma at either room temperature or body temperature was less than 8 ng/cm(2), suggesting that the surfaces might possess good blood compatibility. In addition, the adhesion and detachment of L929 cells could be tuned, and the ability to control the detachment of cells thermally was restored by block polymerization of hydrophobic, cell-adhesive PS onto a thicker PNIPAAm layer. In addition to providing a simple and effective design for advanced cell-culture surfaces, these results suggest new biomedical applications for PNIPAAm.

Cell adhesion on chiral surface: The role of protein adsorption

Chirality is one of the basic, unique, and most appealing features of biological molecules; however, many intriguing chiral phenomena in biological world remains insufficiently revealed yet. In this research, we fabricated chiral surfaces by assembling natural chiral amino acids-cysteine of opposite configurations (D- and L-) onto gold surfaces, respectively, and investigated the adhesion of the L929 fibroblast on them. No significant differences were observed in the density of adherent cells under serum-free culture condition; while in serum-containing condition, significantly more cells adhered on the L-Cys assembled surfaces. This phenomenon suggested that serum protein might play an important role in mediating the selective adhesion of cells on chiral surfaces. Hence, we adopted both radiolabeling and surface plasmon resonance (SPR) techniques to monitor protein adsorption onto the above surfaces. The results evidently showed more proteins adsorbed onto surfaces assembled with L-Cys. We propose that the difference in protein adsorption on chiral surfaces as demonstrated in this paper might not only shed light on the ensuing investigation of bio-related chirality phenomena, but also provide a novel strategy for the rational design and fabrication of novel biomaterials and bio-related devices based on chiral effects.

A surface decorated with diblock copolymer for biomolecular conjugation

A simple and attractive method was introduced to construct bioactive surfaces that exhibit non-specific protein resistant properties and high loading capacities for immobilizing various specific biomolecules. These bioactive surfaces may find wide potential biomedical applications.

Adsorption behavior of human serum albumin on ATR crystal studied by in situ ATR/FTIR spectroscopy and two-dimensional correlation analysis

The time-dependent adsorption behavior of human serum albumin (HSA) onto an ATR (ZnSe) crystal was investigated by two-dimensional (2D) correlation analysis and in situ ATR-FTIR spectroscopy following the secondary structural changes in the amide I region. The two major advantages of the generalized 2D correlation spectroscopy were first tested. New extra bands have been resolved by 2D correlation analysis, but they are either artifacts or a result of uncertainty on band position in generalized 2D correlation spectroscopy. The sequence of the intensity variations of the three sub-bands under the amide I band profile deduced from the sequential order rules is contradictory to the experimental observation, which supports our argument on the sequential order rules in generalized 2D correlation spectroscopy (H. Huang, Anal. Chem., 2007, 79, 8281-8292). Subsequent detailed analysis on the in situ ATR-IR spectra shows that the adsorption process of HSA on the ATR (ZnSe) crystal in aqueous solutions can be divided into three stages: no obvious conformational transitions in the first 25 min of adsorption of HSA molecules; large structural rearrangement from α-helix to random coil and short extended chain structures in a fully cooperative way from 25 to 50 min of adsorption; and further slight conformational transformation of short extended chain and turn structures into random coil with no sequential order after 50 min of adsorption.

Time-dependent adsorption behavior of β-lactoglobulin on ZnSe crystal surface studied by 2D correlation ATR/FTIR spectroscopy

The time-dependent adsorption behavior of β-Lactoglobulin (β-Lg) on ATR crystal (ZnSe) surface was studied by two-dimensional (2D) correlation ATR/FTIR spectroscopy. More bands were resolved by 2D correlation spectroscopy compared to the results from second derivative (SD) and Fourier self-deconvolution (FSD) analyses, but some of the new bands resolved may originate from bandwidth changes, wavenumber shifts, etc. The integrated/overall sequential order of the intensity changes of the four sub-bands in amide I region obtained from 2D correlation spectroscopy was not consistent with the experimental observation. Adsorption-induced conformational changes did not occur until 10 min of adsorption of β-Lg molecules on the ZnSe crystal surface. The relative contents of the low-wavenumber component of the antiparallel β-strands (1627cm(-1)) and random segments with α-helix (1651cm(-1)) changed prior to β-turns (1666cm(-1)) and the high-wavenumber component of the antiparallel β-strands (1684cm(-1)). More specifically, from about 10 to 15min of adsorption, the loss content of the low-wavenumber component of the antiparallel β-strands (1627cm(-1)) was simultaneously transformed into random segments (1651cm(-1)). After 20 min of adsorption, the content of β-turns (1666cm(-1)) started to decrease, and the loss of β-turns (1666cm(-1)) was also transformed into antiparallel β-strands (high-wavenumber component at 1684cm(-1)) in a cooperative way as the β-Lg molecules become more extended.

Conformational Changes of Protein Adsorbed on Tailored Flat Substrates with Different Chemistries

Changes in the bioactivity of a protein after being adsorbed on a material surface may result from conformational changes of the protein. Unfortunately, however, direct evidence of such conformational changes of proteins adsorbed on a flat material surface is sparse so far. This is because probing the conformation of an adsorbed protein on material surfaces, especially flat ones, remains a challenge due to considerable experimental difficulties. In this study, the surface-enhanced Raman scattering (SERS) technique is used to characterize the conformational changes of a protein (lysozyme) adsorbed on tailored flat gold substrates with different chemistries. Two such substrates are formed by self-assembly of octadecanethiol and thiolated PEG on gold chips (Au-C18 and Au-PEG). Preliminary results reveal that, compared to the hydrophobic Au-C18 surface, the hydrophilic Au-PEG surface has much smaller effect on the conformation of lysozyme in aqueous solution, which thereby keeps its high bioactivity. The conformational changes of lysozyme adsorbed on material surfaces with different chemistries are well correlated with changes in its bioactivity.

Altered enzymatic activity of lysozymes bound to variously sulfated chitosans

The purpose of this research is to investigate the effects of the variously sulfated chitosans on lysozyme activity and structure. It was shown that the specific enzymatic activity of lysozyme remained almost similar to the native protein after being bound to 6-O-sulfated chitosan (6S-chitosan) and 3,6-O-sulfated chitosan (3,6S-chitosan), but decreased greatly after being bound to 2-N-6-O-sulfated chitosan (2,6S-chitosan). Meanwhile, among these sulfated chitosans, 2,6S-chitosan induced the greatest conformational change in lysozyme as indicated by the fluorescence spectra. These findings demonstrated that when sulfated chitosans of different structures bind to lysozyme, lysozyme undergoes conformational change of different magnitudes, which results in corresponding levels of lysozyme activity. Further study on the interaction of sulfated chitosans with lysozyme by surface plasmon resonance (SPR) suggested that their affinities might be determined by their molecular structures.

In situ FTIR and generalized 2D IR correlation spectroscopic studies on the crystallization behavior of solution-cast PHB film

AbstractThe crystallization behavior of biosynthesized poly(3-hydroxybutyrate) film cast from 1,1,2,2-tetrachloromethane was studied by in situ FTIR spectroscopy and two-dimensional (2D) correlation analysis. Time-dependent in situ FTIR spectral variations in the C=O stretching region (1,780–1,700xa0cm-1) were monitored and analyzed by a series of data processing methods, including calculation of difference spectrum and second derivative spectrum, Fourier self-deconvolution, curve-fitting and 2D correlation analysis. Four bands have been resolved from the 2D correlation analysis, and the following overall sequential order among the intensity changes of the four bands has been obtained at 1,750u2009>u20091,739u2009>u20091,722u2009>u20091,715xa0cm-1. Combining with the other data processing methods, a curve-fitting approach has been employed to reveal that there are probably five component bands under the C=O band profile, centered at 1,746; 1,737; 1,728; 1,722; and 1,712xa0cm-1. Detailed analysis on the in situ component band intensity variations in the C=O stretching region indicates that the crystalline and amorphous band intensities change simultaneously during the crystallization process, with no local sequential order. Further analysis on the relative area percentage changes of the five component bands suggests that the crystalline component only changes in a fully cooperative manner with part of the amorphous component at the initial crystallization period.n Band area change of each state with crystallization time