Secondary Structure of Organophosphorus Hydrolase in Solution and in Langmuir−Blodgett Film Studied by Circular Dichroism Spectroscopy

Abstract

The secondary structure of organophosphorus hydrolase (OPH) has been studied with circular dichroism (CD) spectroscopy in the far-UV region. The effect of pH on the secondary structure of OPH solution was examined over the pH range from 3.56 to 9.60. As shown on the CD spectra, the secondary structure of OPH is well defined when the pH value is near the isoelectric point (7.6); however, it is destroyed when the pH values are increased or decreased further. This is explained by the loss of helical structure. The pH effect on CD spectra contributes to clarify the optimum pH needed to obtain a stable OPH Langmuir film at the airwater interface and its correlation to the secondary structure of the enzyme. Comparative study of the thermal treatment on the secondary structure of OPH in solution, Langmuir-Blodgett film, and dry film shows that the molecular arrangement plays a dominant role in the thermal stability of OPH. With use of the CDPro software package a quantitative estimation of the secondary structure from the CD spectra of OPH solution was obtained. Results show that there is a decrease in the percentage of the R-helical and an increase of β-strands with the change of pH or temperature.

Enzymes immobilized in Langmuir-Blodgett films: Why determining the surface properties in Langmuir monolayer is important?

“…This strategy was applied for the deposition of biocatalysts with active layers of penicillin G acylase (PGA) (Pastorino et al 2002) which preserved an enzyme widely used for medicine production. Also, we can cite: (iv) galactose oxidase Langmuir-Blodgett (LB) films of poly(3-hexyl thiophene) (P3HT)/stearic acid (SA) for the estimation of lactose in milk and its products to prevent "lactose intolerance" (Zheng et al 2004) where the stability was observed for 120 days; (v) laccase and tyrosinase mixed with octadecyltrimethylammonium bromide and with linoleic acid to detect cathecol (Cabaj et al 2010); (vi) cholesterol oxidase immobilized in LB films consisting of positively charged octadecyltrimethylammonium and nano-sized Prussian-blue clusters (Ohnuki et al 2009); (vii) a uricase-chitosan -n-nonadecanoic acid LB films to detect uric acid ; (viii) Hyaluronidase-phospholipid LB films designed to determine hyaluronic acid by using fluorescence spectroscopy (Monteiro et al 2011); (ix) tyrosinase incorporated in a lipidic layer and the use of lutetium bisphthalocyanine as an electron mediator for the voltammetric detection of phenol derivatives, which include one monophenol (vanillic acid), two diphenols (catechol and caffeic acid) and two triphenols (gallic acid and pyrogallol) (Apetrei et al 2012) (x) a bioelectronic tongue based on lipidic nanostructured layers containing phenol oxidases for the analysis of grapes by means of amperometric and capacitance measurements (Pavinatto et al 2011); and (xi) malate dehydrogenase-cationic lipid systems to determine malic acid (Gur et al 2016). Table I summarizes some highlights on the development of bionsensors based on LB films reported in the last three decades.…”

“…This was, therefore, an interesting study on the correlation of thermodynamic, structural and morphological properties of Gibbs monolayers of enzymes and their deposition to solid supports as LB films. Finally, organophosphorus hydrolase could be investigated in both at the air-water interface and on LB films by using circular dichroism and epifluorescence where the dependence of the structure on the temperature could be evaluated (Zheng et al 2004). These studies allowed for the better detection of paraoxon.…”

Moderate conformational impact of citrate on ovotransferrin considerably increases its capacity to self-assemble at the interface

“…It is known that, when a protein essentially existed in the a-helical conformation, two negatives bands at $209 nm and $222 nm, and a positive band at $190 nm are observed [46][47][48]. On the other hand, for the existence of a b-sheet, a positive and negative band appeared at 190 and 215 nm, respectively [49,50].…”

Effect of metal ions on the secondary structure and activity of calf intestine phosphatase

“…Our research focuses on the substitution of cobalt ions for metal ions starting from native CIP. To explore the conformational changes of CIP during the course of substitution, we employed circular dichroism (CD), a well-known spectroscopic technique for obtaining information on the secondary structure of proteins (14,15), together with methods (SELCON3, CONTIN, CDSSTR) for estimating protein secondary structures from CD spectra (16,17,18,19). The assignments obtained from these methods gave six types of secondary structures: regular α-helix, αR; distorted α-helix, αD; regular β-strand, βR; distorted β-strand, βD; turns, T; and unordered, U.…”