Human Growth Hormone Adsorption Kinetics and Conformation on Self-Assembled Monolayers

Abstract

The adsorption process of the recombinant human growth hormone on organic films, created by self-assembly of octadecyltrichlorosilane, arachidic acid, and dipalmitoylphosphatidylcholine, is investigated and compared to adsorption on silica and methylated silica substrates. Information on the adsorption process of human growth hormone (hGH) is obtained by using total internal reflection fluorescence (TIRF). The intensity, spectra, and quenching of the intrinsic fluorescence emitted by the growth hormone's single tryptophan are monitored and related to adsorption kinetics and protein conformation. For the various alkylated hydrophobic surfaces with differences in surface density and conformational freedom it is observed that the adsorbed amount of growth hormone is relatively large if the alkyl chains are in an ordered structure while the amounts adsorbed are considerably lower for adsorption onto less ordered alkyl chains of fatty acid and phospholipid layers. Adsorption on methylated surfaces results in a relatively large conformational change in the growth hormone's structure, as displayed by a 7 nm blue shift in emission wavelength and a large increase in the effectiveness of fluorescence quenching. Conformational changes are less evident for hGH adsorption onto the fatty acid and phospholipid alkyl chains. Adsorption kinetics on the hydrophilic head groups of the self-assembled monolayers are similar to those on solid hydrophilic surfaces. The relatively small conformational changes in the hGH structure observed for adsorption on silica are even further reduced for adsorption on fatty acid head groups.

Tobacco mosaic virus adsorption on self-assembled and Langmuir–Blodgett monolayers studied by TIRF and SFM

“…The contact angles of the OTS and EDA surfaces are in good agreement with literature values [14,16]. The contact angles attained for the phospholipid headgroups, however, are rather high, a possible result from monolayer rearrangement in the presence of the captive air bubble [13]. The higher contact angle for cationic Et-DPPC monolayers versus the DPPS and DPPC monolayers is attributed to a less dense packing due to steric hindrance and charge repulsion.…”

“…In a recent study we demonstrated that LB films with greater mobility adsorbed less human growth hormone than surfaces of similar chemistry but decreased mobility [13]. We attributed this effect to an unfavorable loss of surface conformational entropy upon protein adsorption.…”

“…Octadecyltrichlorosilane (OTS) and LB films were prepared on fused silica as previously described [12,13]. Silica slides were aminated using N-(2-aminoethyl)(3-aminopropyl)trimethoxysilane (EDA) according to the method of Stenger et al [14] and stored under nitrogen until use.…”

Positioning protein molecules on surfaces: A nanoengineering approach to supramolecular chemistry

“…Nanostructures of thiol molecules serve as templates for attaching proteins. Selectivity of protein adsorption can be achieved by using the variation in protein affinity toward different SAMs (52)(53)(54)(55)(56)(57)(58).…”

Relating material surface heterogeneity to protein adsorption: the effect of annealing of micro-contact-printed OTS patterns

“…Since each of the two fluorescence contributions to the total fluorescence intensity can be separately determined in experiments and the concentration of the af488-HSA solution, c Sol , is known, the surface concentration of surface bound af488-HSA, Γ HSA , can, in principle, be calculated by finding the optical relationship between the two fluorescence components, I Ads and I Sol . The conversion from I Ads and I Sol to Γ HSA used in this study paralleled the quantification of protein adsorbed amount in total internal fluorescence microscopy [30,31] and will not be described in detail here (data not shown). The error in the measurement of the adsorbed HSA surface density was up to ±3.5% (estimated from the standard deviations of protein fluorescence intensity measured within OTS bands).…”