Question

1a.) Collagen is a major protein in connective tissue and the extracellular matrix. Both the Bradford assay and absorbance at 280 nm cannot be used to determine the concentration of collagen. Why is this? No more than two or three sentences, please

1b.) The old way of determining the amino acid composition of a protein involved hydrolyzing the peptide bonds joining the amino acids, separating the amino acids by chromatography, then determining the relative amounts of each. How would a modern scientist determine the amino acid composition of a protein? No more than two or three sentences, please

Answer 1

A- Collagen is a protein which is rich in proline amino acid and very poor in the aromatic amino acid. Aromatic amino acids give the absorbance at 280nm. For measurement of protein concentration by Bradford reagent absorbance of aromatic amino acid is used. So we cannot use Bradford assay for collagen protein estimation.

B- Modern scientist use Mass spectrometry to determine the amino acid composition of a protein. Test protein is ionized which leads to its degradation. Then the fragments are separated based on their mass to charge ratio and then detected.