Comprehensive protein sequence coverage using MagReSyn® Trypsin & MagReSyn® Chymotrypsin

نویسندگان

  • Stoyan Stoychev
  • Isak Gerber
  • Justin Jordaan
چکیده

The application note provides a rapid protocol and adaptable guideline for in-depth characterization of protein sequence. Digestions were performed on a four-protein mix using magnetic microparticles with immobilized proteolytic enzymes, MagReSyn® Trypsin and MagReSyn® Chymotrypsin. The orthogonal cleavage sites of Trypsin and Chymotrypsin provided complementary sets of peptides that resulted in increased sequence coverage of cumulative peptide maps. Furthermore, the digestions were rapid (1 hr, Trypsin; 0.5 hr Chymotrypsin), highly efficient, and reproducible. The method was also adapted for use with the Thermo KingfisherTM Duo magnetic microparticle handling station, in order to fully automate the sample preparation protocol for increased reproducibility in routine operation.

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

منابع مشابه

Toward the complete membrane proteome: high coverage of integral membrane proteins through transmembrane peptide detection.

To attain a comprehensive membrane proteome of two strains of Corynebacterium glutamicum (l-lysine-producing and the characterized model strains), both sample pretreatment and analysis methods were optimized. Isolated bacterial membranes were digested with trypsin/cyanogen bromide or trypsin/chymotrypsin, and a complementary protein set was identified using the multidimensional protein identifi...

متن کامل

Enhanced sequence coverage of proteins in human cerebrospinal fluid using multiple enzymatic digestion and linear ion trap LC-MS/MS.

The cerebrospinal fluid (CSF) provides a ready access into the health state of the central nervous system, and alterations in some CSF proteins have been documented in brain disease. However, the complete variety of proteins is not known and methods to identify protein components are still being developed. The goal of this study was to examine the sequence coverage obtained from human CSF diges...

متن کامل

A combination of neutral loss and targeted product ion scanning with two enzymatic digestions facilitates the comprehensive mapping of phosphorylation sites.

We propose here a new strategy for the exhaustive mapping of phosphorylation sites in the Xenopus laevis Cdc25 phosphatase, which regulates cell cycle progression in eukaryotic cells. Two different MS analyses in a linear IT were used to identify the phosphorylated residues. First, a data-dependent neutral loss (DDNL) analysis triggered the fragmentation of peptides that show enhanced neutral l...

متن کامل

Comparison of the electron capture dissociation fragmentation behavior of doubly and triply protonated peptides from trypsin, Glu-C, and chymotrypsin digestion.

In bottom-up proteomics, proteolytically derived peptides from proteins of interest are analyzed to provide sequence information for protein identification and characterization. Electron capture dissociation (ECD), which provides more random cleavages compared to "slow heating" techniques such as collisional activation, can result in greater sequence coverage for peptides and proteins. Most bot...

متن کامل

Toward the Complete Membrane Proteome HIGH COVERAGE OF INTEGRAL MEMBRANE PROTEINS THROUGH TRANSMEMBRANE PEPTIDE DETECTION*□S

To attain a comprehensive membrane proteome of two strains of Corynebacterium glutamicum (L-lysine-producing and the characterized model strains), both sample pretreatment and analysis methods were optimized. Isolated bacterial membranes were digested with trypsin/ cyanogen bromide or trypsin/chymotrypsin, and a complementary protein set was identified using the multidimensional protein identif...

متن کامل

ذخیره در منابع من


  با ذخیره ی این منبع در منابع من، دسترسی به آن را برای استفاده های بعدی آسان تر کنید

برای دانلود متن کامل این مقاله و بیش از 32 میلیون مقاله دیگر ابتدا ثبت نام کنید

ثبت نام

اگر عضو سایت هستید لطفا وارد حساب کاربری خود شوید

عنوان ژورنال:

دوره   شماره 

صفحات  -

تاریخ انتشار 2016