Linus Pauling: There was a man in England, now at MIT, named Ingram, who determined the detailed
nature of the abnormality in the polypeptide chains by a method that he invented:
paper, two-dimensional electrophoresis and chromatography of peptides. He attacked
the polypeptide chains with trypsin, a proteolytic enzyme which splits each of them
into thirteen short pieces of chain, about deca-peptides, perhaps. And a drop of that
stuff was put on the paper, and the electric field causes it to move from the right
to left and solvent flows and you get a chromatographic separation vertically.
Next slide. There’s only one spot that has shifted when you go from normal hemoglobin
to sickle cell hemoglobin. It’s in that vertical line, not quite at the right edge,
and has moved over from the line a little to the left in the right-hand picture. That
turns out to be the first peptide in the beta chain, and analysis of the peptide shows
that - next slide please - this shows what we see here.
The alpha chain starts valine, leucine, serine, and goes on. The beta chain valine,
histidine, leucine, threonine, proline, glutamate. And the sickle cell beta is valine,
histidine, leucine, threonine, proline, and then valine in the sixth position. The
other 140 positions are occupied by the same amino acid. This involves just one nucleotide
in the gene replaced in such a way that instead of glutamic acid, valine is introduced.
Glutamic acid is really glutamate - the carboxylic group is ionized COO-. Valine has a hydrocarbon side chain, so you’ve lost one negative charge. There are
two beta chains in the molecule. That means you’ve lost two negative charges in going
to sickle cell hemoglobin.