Protein Function (hemoglobin)

Question 1

A researcher has isolated a protein that appears to be involved in synthesis of biofuels in algae. It eludes from a gel filtration column with an apparent native mass of 120 kDa (based on elation of comparable standard proteins), while SDS-PAGE analysis in the presence of mercaptoethanol revels two protein bands of similar intensity corresponding 25 and 35 kDa. Based on this information, this protein is most likely to be:
A)a homotetramer
B)a heterotetramer
C)a homodimer
D)a heterdimer
E)cross-linked by disulfide bonds in its native form.

Answer 1

B)a heterotetramer
Since the protein appears to have equal amounts of 25 kDa and 35 kDa subunits, with a total MW of 120,000, it is most likely a heterotetramer composed of two 25 kDa and two 35 kDa subunits. SDS-PAGE in the presence of the reducing agent mercaptoethanol would reduce (break) any disulfide cross links between or within subunits, but the data do not allow you to conclude whether these are present in the native protein: many intracellular multi-subunits proteins, such as hemoglobin, do not require disulfide bonds to retain their quaternary structure.

Question 2

You have discovered a new form of myoglobin from cockroaches that has a P50 of 40 torr. Whale myoglobin has a P50 of 3 torr. Which of the following statements is true?
A) at a pO2 of 500 torr, cockroach myoglobin is saturated with oxygen, but whale myoglobin is not.
B)at a pO2 of 40 torr, cockroach myoglobin is nearly saturated with oxygen.
C)at pO2 of 3 torr, cockroach myoglobin has very little oxygen bound.
D)whale myoglobin is always less saturated with oxygen than cockroach myoglobin, no matter what the partial pressure of oxygen (pO29 is.
E)cockroach myoglobin must have a sigmoidal binding curve

Answer 2

C)at pO2 of 3 torr, cockroach myoglobin has very little oxygen bound.

Question 3

The binding of one O2 to a molecule of hemoglobin normally results in …….
A)the release of any other O2 that may have bound earlier
B)a decrease in hemoglobins ability to bind a second O2
C)an increased affinity for O2 in the remaining subunits
D)dissociation of heme prosthetic group
E)dissociation of the hemoglobin

Answer 3

C)an increased affinity for O2 in the remaining subunits
The sequential binding of O2 to each subunit in hemoglobin tetramer shifts the Tense (T)- relaxed (R) state equilibrium towards the R state (i.e more like myoglobin), increase the affinity from subsequent O2 binding to the remaining subunits. This is one example of a cooperative phenomenon.

Question 4

In the figure below, the three curves represent hemoglobin binding at three different pH values, pH 7.2, pH 7.4 and pH 7.6. Which of the below statements is true?
A)low pH favors the high affinity binding state
B)low pH favors the low affinity oxygen binding state
C)oxygen affinity is independent of pH
D)oxygen binding is a non- cooperative at low pH
E)oxygen binding is non- cooperative at high pH.

Answer 4

B) low pH favors the low affinity oxygen binding state
The shift in the oxygen binding affinity curve to the right at lower pH is indicative of a decreased affinity of Hb for O 2 (I.e it requires a higher oxygen concentration to achieve a given degree of saturation). All three curves appear to be sigmoidal, indicating there is a subunit cooperativista at all three pH values.