When human hemoglobin undergoes a mutation, the mutant protein usually does not replace all of the normal HbA in the red blood cells or erythrocytes of the individual. The erythro- cytes contain mixtures of varying amounts of both HbA and the mutant protein depending on the mutation and the individual. Hb 2. Yakima is a mutant human Hb with an Asp-(B99)His mutation. The diagram on the right shows that Hb Yakima was separated by DEAE-cellulose chromatography from HbA with a 0 – 0.1 M linear gradient of NaCl buffered to pH 8.3. Why is chromatog- raphy carried out at pH 8.3? If the isoelectric point of HbA is 6.85, what is the change in total charge caused by the mutation?How does the change in charge explain the chromatography elution profile of the Hb Yakima/HbA mixture? 1,5- -Hb-A Hb -Yakima 0.5 20 40 60 80 100 Fraction number O.D578 nm

Biochemistry
9th Edition
ISBN:9781319114671
Author:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Publisher:Lubert Stryer, Jeremy M. Berg, John L. Tymoczko, Gregory J. Gatto Jr.
Chapter1: Biochemistry: An Evolving Science
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When human hemoglobin undergoes a mutation, the
mutant protein usually does not replace all of the normal HbA in
the red blood cells or erythrocytes of the individual. The erythro-
cytes contain mixtures of varying amounts of both HbA and the
mutant protein depending on the mutation and the individual. Hb
Yakima is a mutant human Hb with an Asp-(B99)His mutation.
The diagram on the right shows that Hb Yakima was separated
by DEAE-cellulose chromatography from HbA with a 0 – 0.1 M
linear gradient of NaCl buffered to pH 8.3. Why is chromatog-
raphy carried out at pH 8.3? If the isoelectric point of HbA is 6.85,
what is the change in total charge caused by the mutation?How
does the change in charge explain the chromatography elution
profile of the Hb Yakima/HbA mixture?
1,5
-Hb-A
Hb -Yakima
1.0
0.5-
20
40
60
80
00
Fraction number
O.D578 nm
Transcribed Image Text:When human hemoglobin undergoes a mutation, the mutant protein usually does not replace all of the normal HbA in the red blood cells or erythrocytes of the individual. The erythro- cytes contain mixtures of varying amounts of both HbA and the mutant protein depending on the mutation and the individual. Hb Yakima is a mutant human Hb with an Asp-(B99)His mutation. The diagram on the right shows that Hb Yakima was separated by DEAE-cellulose chromatography from HbA with a 0 – 0.1 M linear gradient of NaCl buffered to pH 8.3. Why is chromatog- raphy carried out at pH 8.3? If the isoelectric point of HbA is 6.85, what is the change in total charge caused by the mutation?How does the change in charge explain the chromatography elution profile of the Hb Yakima/HbA mixture? 1,5 -Hb-A Hb -Yakima 1.0 0.5- 20 40 60 80 00 Fraction number O.D578 nm
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