TY - JOUR

T1 - Search and Identification of Amyloid Proteins

AU - Belashova , Tatyana A.

AU - Valina, Anna A.

AU - Sysoev , Evgeniy I.

AU - Velizhanina , Maria E.

AU - Zelinsky, Andrew A.

AU - Galkin, Alexey P.

N1 - Belashova, T.A.; Valina, A.A.; Sysoev, E.I.; Velizhanina, M.E.; Zelinsky, A.A.; Galkin, A.P. Search and Identification of Amyloid Proteins. Methods Protoc. 2023, 6, 16. https://doi.org/10.3390/mps6010016

PY - 2023/2/4

Y1 - 2023/2/4

N2 - Amyloids are fibrillar proteins with a cross-β structure. Pathological amyloids are associated with the development of a number of incurable diseases, while functional amyloids regulate vital processes. The detection of unknown amyloids in living objects is a difficult task, and therefore the question of the prevalence and biological significance of amyloids remains open. We present a description of two methods, the combination of which makes it possible to find and identify amyloid proteins in the proteome of various organisms. The method of proteomic screening for amyloids allows the detection of the proteins that form SDS-resistant aggregates. SDS resistance is a general feature of amyloid fibrils. Protein aggregates resistant to SDS treatment can be collected by ultracentrifugation and further identified by mass spectrometry. However, in addition to amyloids, SDS-resistant aggregates contain some non-amyloid proteins. To test the amyloid properties of proteins identified by proteomic screening, we developed the method of fibril immunoprecipitation followed by Congo red staining and birefringence analysis. The methods of proteomic screening and immunoprecipitation of fibrillar proteins have been successfully tested and applied for the identification of amyloid proteins in yeast and vertebrates.

AB - Amyloids are fibrillar proteins with a cross-β structure. Pathological amyloids are associated with the development of a number of incurable diseases, while functional amyloids regulate vital processes. The detection of unknown amyloids in living objects is a difficult task, and therefore the question of the prevalence and biological significance of amyloids remains open. We present a description of two methods, the combination of which makes it possible to find and identify amyloid proteins in the proteome of various organisms. The method of proteomic screening for amyloids allows the detection of the proteins that form SDS-resistant aggregates. SDS resistance is a general feature of amyloid fibrils. Protein aggregates resistant to SDS treatment can be collected by ultracentrifugation and further identified by mass spectrometry. However, in addition to amyloids, SDS-resistant aggregates contain some non-amyloid proteins. To test the amyloid properties of proteins identified by proteomic screening, we developed the method of fibril immunoprecipitation followed by Congo red staining and birefringence analysis. The methods of proteomic screening and immunoprecipitation of fibrillar proteins have been successfully tested and applied for the identification of amyloid proteins in yeast and vertebrates.

KW - amyloids

KW - prions

KW - Proteomic screening

KW - fibril immunoprecipitation

KW - detergent resistance

KW - mass spectrometry

KW - electron microscopy

KW - Congo red

UR - https://www.mdpi.com/2409-9279/6/1/16

M3 - Review article

VL - 6

JO - Methods and Protocols — Open Access Journal

JF - Methods and Protocols — Open Access Journal

SN - 2409-9279

IS - 1

M1 - 16

ER -