Analysis of the amino acid sequences of plant Bowman-Birk inhibitors. Selvaraj M. Murthy Y.
Analysis of the amino acid sequences of plant Bowman-Birk inhibitors
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N- and C-Terminal Amino Acid Analysis
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Note that we determine asparagine as aspartic acid, and glutamine as glutamic acid. We perform the analysis on a BioChrom 30 amino acid analyser. It makes ion-exchange chromatography with post-column derivatization using ninhydrin. We can thus monitor all amino acids except Proline nm at nm. First, we place the sample in a 0.
We then place the tube in a vessel with an acid hydrolysis solution 6M HCl containing, 0.
The vessel is flushed with argon. After hydrolysis the test sample is dried in vacuum. Next, we dissolve the sample in loading buffer sodium citrate pH 2. We use this as an internal standard to correct for changes in reagent stability and flow rates.go site
Amino Acid Sequence and Terminal Sequence Analysis
We analyze a standard mixture of amino acids at 3 different concentrations levels double determination. This shows the linearity within the measurement range to pmol. Next, a plot shows the peak area versus the known concentration of each standard solution. It passes when the r-value of the calibration is better than 0.
Depending on the amino acid residue, the signal goes into saturation around nmol. We use the results to determine the precision and the accuracy of the test. A standard amino acid solution is analyzed for each 5 samples.
Analysis of the amino acid sequences of plant Bowman-Birk inhibitors | SpringerLink
Based on at least two runs, a we then calculate a compensation factor to correct for ninhydrin activity. We correct each amino acid in the sample by this compensation factor. Known protein samples : If we know the protein sequence, we then divide the measured AA amount in pmol by the number of each specific amino acid in the sequence providing a pmol protein. The calculations against known amino acid sequence is then made. We use a best linear fit based on all detected amino acids. It is modified for each experiment not to include amino acids with greater than 5 per cent variation.
We can analyze most samples that contain proteins and peptides. Avoid very high salt concentrations. Do not use buffer that contain high amounts of primary amines, e. Avoid fingerprints and dust in the sample. These contain large amounts of human proteins that will obscure the results. Solid material should be provided in sufficient amount for accurate weighing of samples at our facilities mg material. Put the sample into the microcentrifuge tube. Lyophilize or speed vac the protein to ensure protein stability during shipment. Remember to print and include the sample submission form. Then follow these simple instructions:.