caa a22 4500 445298642 CHVBK 20180317142550.0 cr unu---uuuuu 170323e20100401xx s 000 0 eng 10.1007/s12013-009-9073-4 doi (NATIONALLICENCE)springer-10.1007/s12013-009-9073-4 Langmuir Films from Human Placental Membranes: Preparation, Rheology, Transfer to Alkylated Glasses, and Sigmoidal Kinetics of Alkaline Phosphatase in the Resultant Langmuir-Blodgett Film [Elektronische Daten] [Eduardo Clop, María Perillo] In the present study, we studied the activity of human placental alkaline phosphatase (PLAP) constraint in a planar surface in controlled molecular packing conditions. For the first time, Langmuir films (LFs) were prepared by the spreading of purified placental membranes (PPM) on the air-water interface and their stability and rheological properties were studied. LFs exhibited a collapse pressure πC=48mN/m, hysteresis during the compression-decompression cycle (C-D), indicating a plastic deformation, and a compressibility modulus (K) compatible with liquid-expanded phases. A phase transition point appeared at πT=28mN/m and, following successive C-D, it moved toward lower surface areas and higher K, suggesting the lost of some non-PLAP proteins as components of vesicles that might protrude from the monolayer (confirmed by combining lipid/protein molar ratio analysis, PAGE-SDS and V max). LFs were transferred at 35mN/m to alkylated glasses to obtain Langmuir-Blodgett films (LB35) the stability of which was confirmed by AFM. The kinetics of p-nitrophenyl phosphate (pNPP) hydrolysis at 37°C catalyzed by PPM was Michaelian and exhibited the thermostability at 60°C typical of PLAP. In LB35, PLAP exhibited a sigmoidal kinetics which resembled the behavior of the partially metalated enzyme but might become from a cross-talk between protein and membrane structures. Springer Science+Business Media, LLC, 2009 Human placental membranes nationallicence Alkaline phosphatase nationallicence Langmuir-Blodgett films nationallicence p NPP hydrolysis nationallicence Sigmoidal kinetics nationallicence A1, A2, and A3 : Monolayer aspirated at 35mN/m during the 1st, 2nd, and 3rd C-D cycles, respectively nationallicence AFM : Atomic force microscopy nationallicence AP : Alkaline phosphatase nationallicence C-D : Compression-decompression cycle nationallicence % CV : Percent variation coefficient nationallicence EDTA : Ethylendiamintetraacetic acid nationallicence GPI : Glycophostatidyl-inositol nationallicence K M : Michaelis-Menten constant nationallicence LB35 : Langmuir-Blodgett film packed at 35mN/m nationallicence LF : Langmuir film nationallicence 2-ME : 2-Mercaptoethanol nationallicence M r : Relative mobility nationallicence MWM : Molecular weight markers nationallicence PAGE : Polyacrylamide gel electrophoresis nationallicence PLAP : Placental alkaline phosphatase nationallicence p NPP : para-nitrophenylphosphate nationallicence p NPx : para-nitro phenoxide nationallicence PPM : Purified placental membrane nationallicence SDS : Sodium dodecyl-sulfate nationallicence s.e.m. : Standard error of the mean nationallicence V max : Maximal velocity nationallicence V 0 : Initial velocity nationallicence Clop Eduardo Química Biológica-Biofísica Química, Dpto. Química, Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 1611, X 5016GCA, Córdoba, Argentina aut Perillo María Química Biológica-Biofísica Química, Dpto. Química, Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 1611, X 5016GCA, Córdoba, Argentina aut Cell Biochemistry and Biophysics Humana Press Inc 56/2-3(2010-04-01), 91-107 1085-9195 56:2-3<91 2010 56 12013 https://doi.org/10.1007/s12013-009-9073-4 text/html Onlinezugriff via DOI 1 research-article jats NATIONALLICENCE 856 40 https://doi.org/10.1007/s12013-009-9073-4 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Clop Eduardo Química Biológica-Biofísica Química, Dpto. Química, Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 1611, X 5016GCA, Córdoba, Argentina aut NATIONALLICENCE 700 1- Perillo María Química Biológica-Biofísica Química, Dpto. Química, Facultad de Ciencias Exactas, Físicas y Naturales, Universidad Nacional de Córdoba, Av. Vélez Sarsfield 1611, X 5016GCA, Córdoba, Argentina aut NATIONALLICENCE 773 0- Cell Biochemistry and Biophysics Humana Press Inc 56/2-3(2010-04-01), 91-107 1085-9195 56:2-3<91 2010 56 12013 Metadata rights reserved Springer special CC-BY-NC licence nationallicence BK010053 XK010053 XK010000 NATIONALLICENCE NATIONALLICENCE NL-springer