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The complete enzymic hydrolysis of crosslinked proteins
Advances in Experimental Medicine and Biology
|January 1, 1977
Summary
Complete enzymic hydrolysis effectively detects protein crosslinks, including isopeptide bonds. However, high crosslinking levels or racemization can hinder detection, and pre-cleavage of disulfide bonds isn't always necessary.
Area of Science:
- Biochemistry
- Proteomics
- Analytical Chemistry
Background:
- Protein crosslinking is crucial for structural integrity.
- Accurate detection of crosslinks is vital for understanding protein function and disease.
- Existing methods for crosslink analysis have limitations.
Purpose of the Study:
- To review procedures for complete enzymic hydrolysis of proteins.
- To evaluate the application of this method for detecting various protein crosslinks.
- To identify limitations and necessary pre-treatments for enzymic hydrolysis in crosslink detection.
Main Methods:
- Comprehensive review of published procedures for complete protein hydrolysis using enzymes.
- Analysis of case studies demonstrating the use of enzymic hydrolysis for isopeptide and chemically induced crosslink detection.
- Discussion of factors affecting method success, such as crosslinking density and stereochemistry.
Main Results:
- Complete enzymic hydrolysis is a successful technique for identifying naturally occurring isopeptide crosslinks.
- The method is also effective for detecting various chemically introduced crosslinks.
- Method failure can occur with excessive crosslinking or associated racemization.
- The necessity of pre-cleaving disulfide bonds before enzymic digestion is questioned.
Conclusions:
- Complete enzymic hydrolysis offers a valuable approach for protein crosslink analysis.
- Understanding the limitations related to crosslinking extent and stereochemistry is important for reliable results.
- Re-evaluation of disulfide bond pre-treatment protocols may be warranted for specific applications.