Modification of C Terminus Provides New Insights into the Mechanism of alpha-Synuclein Aggregation

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388963%3A_____%2F17%3A00485440" target="_blank" >RIV/61388963:_____/17:00485440 - isvavai.cz</a>

Výsledek na webu

<a href="http://dx.doi.org/10.1016/j.bpj.2017.08.027" target="_blank" >http://dx.doi.org/10.1016/j.bpj.2017.08.027</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1016/j.bpj.2017.08.027" target="_blank" >10.1016/j.bpj.2017.08.027</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Modification of C Terminus Provides New Insights into the Mechanism of alpha-Synuclein Aggregation

Popis výsledku v původním jazyce

Aggregation of neuronal protein alpha-synuclein leads to the formation of amyloid fibrils, which are associated with the development of Parkinson's disease. The mechanism of alpha-synuclein pathology is not fully understood and is a subject of active research in the field. To tackle this problem, the fusions of fluorescent proteins to alpha-synuclein C-terminus are often used in cellular and animal studies. The effects induced by such alpha-synuclein sequence extension on alpha-synuclein aggregation propensity are, however, not systematically examined despite the evidence that the negatively charged C-terminus plays a critical role in the regulation of alpha-synuclein aggregation. In this work, we investigated how the charge and length variations of the C-terminus affect the aggregation propensity of alpha-synuclein. To address these questions, we prepared mutants of alpha-synuclein carrying additional moieties of different charge and length at the protein C-terminus. We determined the rates of two different aggregation stages (primary nucleation and elongation) based on a thioflavin T kinetic assay. We observed that all mutants bearing neutrally charged moieties of different length fibrilized slower than wild-type alpha-synuclein. The primary nucleation and elongation rates strongly decreased with increase of the C-terminal extension length. Meanwhile, charge variation of the C-terminus significantly changed the rate of alpha-synuclein nucleation, but did not markedly affect the rate of fibril elongation. Our data demonstrate that both the charge and length of the C-terminus play an important role at the stage of initial fibril formation, but the stage of fibril elongation is affected mainly by the length of C-terminal extension. In addition, our results suggest that there are at least two steps of incorporation of alpha-synuclein monomers into the amyloid fibril: namely, the initial monomer binding to the fibril end (charge-dependent, relatively fast), and the subsequent conformational change of the protein (charge-independent, relatively slow, and thus the rate-limiting step).

Název v anglickém jazyce

Modification of C Terminus Provides New Insights into the Mechanism of alpha-Synuclein Aggregation

Popis výsledku anglicky

Aggregation of neuronal protein alpha-synuclein leads to the formation of amyloid fibrils, which are associated with the development of Parkinson's disease. The mechanism of alpha-synuclein pathology is not fully understood and is a subject of active research in the field. To tackle this problem, the fusions of fluorescent proteins to alpha-synuclein C-terminus are often used in cellular and animal studies. The effects induced by such alpha-synuclein sequence extension on alpha-synuclein aggregation propensity are, however, not systematically examined despite the evidence that the negatively charged C-terminus plays a critical role in the regulation of alpha-synuclein aggregation. In this work, we investigated how the charge and length variations of the C-terminus affect the aggregation propensity of alpha-synuclein. To address these questions, we prepared mutants of alpha-synuclein carrying additional moieties of different charge and length at the protein C-terminus. We determined the rates of two different aggregation stages (primary nucleation and elongation) based on a thioflavin T kinetic assay. We observed that all mutants bearing neutrally charged moieties of different length fibrilized slower than wild-type alpha-synuclein. The primary nucleation and elongation rates strongly decreased with increase of the C-terminal extension length. Meanwhile, charge variation of the C-terminus significantly changed the rate of alpha-synuclein nucleation, but did not markedly affect the rate of fibril elongation. Our data demonstrate that both the charge and length of the C-terminus play an important role at the stage of initial fibril formation, but the stage of fibril elongation is affected mainly by the length of C-terminal extension. In addition, our results suggest that there are at least two steps of incorporation of alpha-synuclein monomers into the amyloid fibril: namely, the initial monomer binding to the fibril end (charge-dependent, relatively fast), and the subsequent conformational change of the protein (charge-independent, relatively slow, and thus the rate-limiting step).

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10610 - Biophysics

Projekt

—

Návaznosti

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2017

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Biophysical Journal

ISSN

0006-3495

e-ISSN

—

Svazek periodika

113

Číslo periodika v rámci svazku

10

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

10

Strana od-do

2182-2191

Kód UT WoS článku

000416213100010

EID výsledku v databázi Scopus

2-s2.0-85029605070