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Biochem Contents
Biochemistry Protocols
Amino Acids and Proteins
Protein Purification
Protein Crystallization
Proteolytic Cleavage
Peptide Mapping
Lipids
Lipid Purification
DGK Assay
Liposome Swelling Assay
Lipid Extraction
Seperation and Quantification
Lipid Transmethylation
Enzymes
ATPase Assays
Kinase and Phosphotase Assays
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Proteolytic Cleavage
Peptide bonds between amino acids in proteins can be broken by a process known as proteolytic cleavage. This is carried out by lytic enzymes such as peptidases, proteases or proteolytic cleavage enzymes.
Naturally, proteins undergo proteolytic processing as a form of structural integrity conservation. This may be to counter misfolding due to environmental conditions or other functional mechanisms such as correcting structure in protein secretion or membrane implantation. Taking advantage of this mechanism, it is possible to to work with specific peptide fragements rather than entire proteins.
Partial Proteolytic Cleavage
In-Situ Proteolysis
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Peptide Mapping
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Peptide mapping involves controlled cleavage of a pure protein with small amounts of a pure protease to generate peptides of characteristic, reproducible sizes. These peptides can be separated on PAGE to produce a â€fingerprint†characteristic of the protein. Peptide mapping can map cleavage sites in an unknown protein, or it can identify an unknown protein based upon its fingerprint identity with a previously tested sample.
The polyacrylamide gel used can be either denaturing or non-denaturing, but SDS PAGE is most often used because it gives molecular weight information about the peptides produced. Small amounts of protease are used, so that minor variations in time and temperature of incubations will not overly perturb the results.
Proteins for peptide mapping can be taken from bands sliced out of electrophoresis gels, or purified by standard means. Protocols are provided for the mapping of a pure protein, a protein in an acrylamide gel, and a protein isolated from National Diagnostics' ProtoPrep II Matrix.
Tryptic or Chymotryptic Peptide Mapping
Cynaogen Bromide Peptide Mapping
The Cleveland Method: SDS-PAGE Peptide Mapping
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Lipid Purification
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Column Purification of Demethylated Sphingomyelin
LIPID EXTRACTION
Papers: Membrane Lipids :: isolation & purification
A Method for Micro-Scale Isolation and Purification of Gangliosides
Purification of fatty acids before analysis
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DGK Assay
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DGK Assay
DGK Membrane Preparation
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Liposome Swelling Assay
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Liposome Swelling Assay
Liposome Preparation
Notes
Thin Film Hydration
Lipoplex and LPD Nanoparticles for In Vivo Gene Delivery
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Biochem Contents
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Protein Purification
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