Optimization of Conditions for Blood Plasma Peptidome Analysis


Дәйексөз келтіру

Толық мәтін

Ашық рұқсат Ашық рұқсат
Рұқсат жабық Рұқсат берілді
Рұқсат жабық Тек жазылушылар үшін

Аннотация

The conditions for peptidome analysis of blood plasma were optimized and the efficacy of the proposed approach was compared with the methods described in the literature. The method implies solution of two main problems: inactivation of blood plasma proteases and dissociation of peptides from major blood plasma proteins, which they are quantitatively associated with. To solve these problems, we proposed a new method of sample preparation. The essence of the method is simultaneous denaturation of plasma proteins plus reduction and alkylation of thiol groups of Cys, which is achieved by heating a blood plasma sample (95°C) in the presence of sodium deoxycholate, tris(2-carboxyethyl)phosphine, and 2-chloroacetamide. After separation of peptides from proteins by ultrafiltration on microcentrifuge filters and removal of sodium deoxycholate, the peptides are identified by LC-MS/MS using a Q Exactive HF (Thermo Scientific) mass spectrometer. As a result of one LC-MS/MS run of the peptide mixture obtained from ~15 μL of blood plasma, 2257 peptide fragments of 867 proteins were identified, which is 1.5 times higher than the values achieved by using the generally accepted method of differential solubilization. Our immediate plans include the use of our approach for cataloguing human blood plasma peptides, as well as establishing the magnitude of individual variability and the features of the peptidome that are related to gender and age.

Негізгі сөздер

Авторлар туралы

R. Ziganshin

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry

Хат алмасуға жауапты Автор.
Email: rustam.ziganshin@gmail.com
Ресей, Moscow, 117997

V. Ryabinin

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry

Email: rustam.ziganshin@gmail.com
Ресей, Moscow, 117997

I. Azarkin

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry

Email: rustam.ziganshin@gmail.com
Ресей, Moscow, 117997

V. Govorun

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry; Federal Research and Clinical Center of Physical–Chemical Medicine

Email: rustam.ziganshin@gmail.com
Ресей, Moscow, 117997; Moscow, 119435

V. Ivanov

Shemyakin–Ovchinnikov Institute of Bioorganic Chemistry

Email: rustam.ziganshin@gmail.com
Ресей, Moscow, 117997


© Pleiades Publishing, Ltd., 2018

Осы сайт cookie-файлдарды пайдаланады

Біздің сайтты пайдалануды жалғастыра отырып, сіз сайттың дұрыс жұмыс істеуін қамтамасыз ететін cookie файлдарын өңдеуге келісім бересіз.< / br>< / br>cookie файлдары туралы< / a>