Protein Sequencing Protocols -- (1) (2)
Protein sequencing - From Wikipedia, the free encyclopedia
Introduction: Proteins are
found in every cell and are essential to every biological process, protein structure is very complex: determining a protein's structure
involves first protein sequencing - determining the amino acid sequences of its constituent peptides; and also determining what conformation
it adopts and whether it is complexed with any non-peptide molecules. Discovering the structures and functions of proteins in living
organisms is an important tool for understanding cellular processes, and allows drugs that target specific metabolic pathways to be
invented more easily.
The two major direct methods of protein sequencing are mass spectrometry and the Edman degradation reaction.
It is also possible to generate an amino acid sequence from the DNA or mRNA sequence encoding the protein, if this is known. However,
there are a number of other reactions which can be used to gain more limited information about protein sequences and can be used as
preliminaries to the aforementioned methods of sequencing or to overcome specific inadequacies within them.
Contents
1 Determining
amino acid composition
2 N-terminal amino acid analysis
3 C-terminal amino
acid analysis
5 The Edman degradation reaction
5.1 Limitations of the Edman degradation
7
Predicting protein sequence from DNA/RNA sequences
Other Protein Sequencing Protocols
SAMPLE PREPARATION TIPS
FOR THE PROTEIN SEQUENCERS
(Additional Protein Sequencing Sample Preparation Tips can be found at: ABRF Home Page )
In-Gel Digest procedure for
obtaining internal peptide sequences from proteins with blocked N-termini.
On-PVDF membrane digest procedure for obtaining internal
peptide sequences from proteins with blocked N-termini.
N-terminal Protein/Peptide Sequencing
Carboxyl-Terminal Sequencing Methods
- Shively, John
Peptide Sequence Databases
nr
All non-redundant GenBank CDS translations+PDB+SwissProt+PIR+PRF
month
All new or revised GenBank CDS translation+PDB+SwissProt+PIR+PRF released in the last 30 days.
swissprot
Last major release
of the SWISS-PROT protein sequence database (no updates)
Drosophila genome
Drosophila genome proteins provided by Celera and BerkeleyDrosophila Genome Project (BDGP).
yeast
Yeast (Saccharomyces cerevisiae) genomic CDS translations
ecoli
Escherichia coli genomic CDS
translations
pdb
Sequences derived from the 3-dimensional structure from Brookhaven Protein Data Bank
kabat [kabatpro]
Kabat's database of
sequences of immunological interest
alu
Translations of select Alu repeats from REPBASE, suitable for masking Alu repeats from query sequences. It is available by anonymous FTP from ncbi.nlm.nih.gov (under the /pub/jmc/alu directory). See "Alu alert" by Claverie and Makalowski, Nature vol. 371, page 752 (1994) .
Sequence Databases & Tools (Keyword Search for Gene/Protein Sequences)