HEAT-STABLE LANGMUIR-BLODGETT-FILM OF GLUTATHIONE-S-TRANSFERASE

2020-04-05 20:20:24

The function film activity enzyme

责任者: ANTOLINI, F;PADDEU, S;NICOLINI, C 单位: UNIV GENOA,SCH MED,INST BIOPHYS,I-16153 GENOA,ITALY.;TECHNOBIOCHIP,I-57030 MARCIANA,ITALY. 来源出处: LANGMUIR, v 11, JUL 1995, p 2719- 2725 摘要: Glutathione-S-tranferase (GST) is an enzyme able to conjugate glutathione to electrophilic molecules such as 1-chloro-2,4-dinitrobenzene or some triazinic pesticides, such as atrazine, with production of protons. The Langmuir-Blodgett (LB) technique was utilized and optimized to obtain stable monolayers of this enzyme. The film was then characterized by means of circular dichroism spectroscopy, nanogravimetry, and spectrophotometry in order to determine the structure, surface density, and functional activity of the enzyme when packed in the monolayers. The functional analysis of the LB film was also conducted as a function of the number of layers and of the temperature. While the kinetic response as a function of layers suggested that only the external layer was active, the film activity as a function of temperature indicated that, as shown earlier for other proteins, GST in a LB film appears to strictly preserve its functional activity and thus its structure up to 423 K. The enzymatic activity was finally tested in solution with the potentiometric alternating biosensor (PAB) system, with the minimum detectable amount of 1 chloro-2,4-dinitrobenzene being 25 mu M. The analysis was highly encouraging for a biosensor application, considering that both structure and function appear largely preserved after the deposition. 关键词: ANISOTROPIC PITTING; CHLORIDE SOLUTIONS; PASSIVITY; PIT INITIATION; PIT MORPHOLOGY; PITTING POTENTIAL MORPHOLOGY; PITTING POTENTIAL; ADDRESSABLE POTENTIOMETRIC SENSOR; ISOENZYME 3-3; BIOELECTRONICS; MUTAGENESIS; CONJUGATION; BIOSENSORS; HERBICIDES; ENZYMES; STEP MUTAGENESIS; CONJUGATION; BIOSENSORS; HERBICIDES; ENZYMES; STEP