Protein synthesis is a central process in all cells, crucial for cell development and maintenance. Translational dysregulation, in fact, is associated with cancer or neurodegenerative diseases. Active protein synthesis occurs on a supramolecular complex, named polyribosome or polysome, formed by a mRNA associated with multiple ribosomes. Polysomes therefore can be considered as a privileged molecular platform to obtain information about the physiological or pathological state in cells. The classical methods for purifying the mRNAs associated with polysomes mainly rely on ultracentrifugation in sucrose gradient followed by standard RNA extraction. This method present several drawbacks, among all it is a time-consuming procedure, which requires a fairly large amounts of starting material. New methods offering an efficient, rapid and user-friendly alternative to standard methods are therefore highly desirable. Here, a panel of surfaces and surface functionalizations were screened for their ability to entrap polysomes with the ultimate aim to set up smart biofunctional surfaces for the purification of nonlabelled polysomes and their associated mRNAs. As a proof-of-concept, prepurified ribosomes and polysomes were incubated on multiple functional surfaces and characterized by atomic force microscopy to assess number and morphology of entrapped polysomes. Surfaces able to efficiently capture polysomes were then included in a microdevice with promising results, opening the future perspective of developing protocols and devices based on biofunctional surfaces.

Biofunctional Surfaces for Smart Entrapment of Polysomes.

Lunelli, L.;Marocchi, L.;Pasquardini, L.;Vanzetti, L.;Viero, G.;Potrich, C.
;
Pederzolli, C.
2021

Abstract

Protein synthesis is a central process in all cells, crucial for cell development and maintenance. Translational dysregulation, in fact, is associated with cancer or neurodegenerative diseases. Active protein synthesis occurs on a supramolecular complex, named polyribosome or polysome, formed by a mRNA associated with multiple ribosomes. Polysomes therefore can be considered as a privileged molecular platform to obtain information about the physiological or pathological state in cells. The classical methods for purifying the mRNAs associated with polysomes mainly rely on ultracentrifugation in sucrose gradient followed by standard RNA extraction. This method present several drawbacks, among all it is a time-consuming procedure, which requires a fairly large amounts of starting material. New methods offering an efficient, rapid and user-friendly alternative to standard methods are therefore highly desirable. Here, a panel of surfaces and surface functionalizations were screened for their ability to entrap polysomes with the ultimate aim to set up smart biofunctional surfaces for the purification of nonlabelled polysomes and their associated mRNAs. As a proof-of-concept, prepurified ribosomes and polysomes were incubated on multiple functional surfaces and characterized by atomic force microscopy to assess number and morphology of entrapped polysomes. Surfaces able to efficiently capture polysomes were then included in a microdevice with promising results, opening the future perspective of developing protocols and devices based on biofunctional surfaces.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11582/324350
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