Adsorption study of alkaline phosphatase in Langmuir monolayers containing cholesterol-rich lipidic microdomains

• Grant number: 15/03594-4
• Support Opportunities: Scholarships in Brazil - Master
• Start date: August 01, 2015
• End date: June 30, 2017
• Field of knowledge: Physical Sciences and Mathematics - Chemistry - Physical-Chemistry
• Agreement: Coordination of Improvement of Higher Education Personnel (CAPES)
• Principal Investigator: Ana Paula Ramos
• Grantee: Rafael Derradi
• Host Institution: Faculdade de Filosofia, Ciências e Letras de Ribeirão Preto (FFCLRP). Universidade de São Paulo (USP). Ribeirão Preto , SP, Brazil

Abstract

Biomineralization is a complex process involving different enzymes, among then the tissue-nonspecific alkaline phosphatase (TNAP). The studies of TNAP inserted in cell membrane model systems, such as proteoliposomes and Langmuir monolayers, are promising and may assist the understanding of such process. The cholesterol in proteic microdomains of biological membranes plays a key role in enzyme activity regulation and biomineralization. The goal of this project is to study the cholesterol effect in the alkaline phosphatase adsorption in biomimetic membrane systems. These systems will be obtained by using Langmuir monolayers in domains in the presence or absence of sphingolipids, a component of "lipid rafts". In this study, Langmuir monolayers will be used to obtain À x A isotherms for different phospholipid, sphingolipid, and cholesterol compositions in the presence and absence of TNAP. The influence of cholesterol and monolayers composition in the formation and morphology of lipid-protein microdomains will be monitored by fluorescence microscopy in a Langmuir trough attached to a fluorescence microscopy, using fluorophore-linked cholesterol and dipalmitoylphosphatidylcholine (DPPC). The systems will also be characterized by dynamic surface tension and rheology measurements using the drop shape method. The advantages of such systems compared to proteoliposomes are the possibility to determine protein/lipid relationship, estimate area per unity of protein, and the activity/pressure relationship. The entire study will be performed under co-supervision of Prof. Pietro Ciancaglini (DQ-FFCLRP). (AU)

Scientific publications

(References retrieved automatically from Web of Science and SciELO through information on FAPESP grants and their corresponding numbers as mentioned in the publications by the authors)

DERRADI, R.; BOLEAN, M.; SIMAO, A. M. S.; CASELI, L.; MILLAN, J. L.; BOTTINI, M.; CIANCAGLINI, P.; RAMOS, A. P.. Cholesterol Regulates the Incorporation and Catalytic Activity of Tissue-Nonspecific Alkaline Phosphatase in DPPC Monolayers. Langmuir, v. 35, n. 47, p. 15232-15241, NOV 26 2019. (17/08892-9, 15/03594-4, 14/11941-3, 16/21236-0)