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13. The Isoelectric Point of Histones Histones are proteins found in eukaryotic cell nuclei, tightly
bound to DNA, which has many phosphate groups. The pI of histones is very high, about 10.8. What
amino acid residues must be present in relatively large numbers in histones? In what way do these
residues contribute to the strong binding of histones to DNA?
Answer Large numbers of positively charged (high pKa) groups in a protein give it a high pI.
In histones, the positively charged R groups of Lys, Arg, and His residues interact strongly
with the negatively charged phosphate groups of DNA through ionic interactions.
14. Solubility of Polypeptides One method for separating polypeptides makes use of their different
solubilities. The solubility of large polypeptides in water depends upon the relative polarity of their R
groups, particularly on the number of ionized groups: the more ionized groups there are, the more soluble
the polypeptide. Which of each pair of the polypeptides that follow is more soluble at the indicated pH?
(a) (Gly)20 or (Glu)20 at pH 7.0
(b) (Lys–Ala)3 or (Phe–Met)3 at pH 7.0
(c) (Ala–Ser–Gly)5 or (Asn–Ser–His)5 at pH 6.0
(d) (Ala–Asp–Gly)5 or (Asn–Ser–His)5 at pH 3.0
Answer
(a) (Glu)20; it is highly negatively charged (polar) at pH 7. (Gly)20 is uncharged except for
the amino- and carboxyl-terminal groups.
(b) (Lys–Ala)3; this is highly positively charged (polar) at pH 7. (Phe–Met)3 is much less
polar and hence less soluble.
(c) (Asn–Ser–His)5; both polymers have polar Ser side chains, but (Asn–Ser–His)5 also has
the polar Asn side chains and partially protonated His side chains.
(d) (Asn–Ser–His)5; at pH 3, the carboxylate groups of Asp residues are partially protonated
and neutral, whereas the imidazole groups of His residues are fully protonated and posi-
tively charged.
15. Purification of an Enzyme A biochemist discovers and purifies a new enzyme, generating the purifi-
cation table below.
(a) From the information given in the table, calculate the specific activity of the enzyme after each
purification procedure.
(b) Which of the purification procedures used for this enzyme is most effective (i.e., gives the greatest
relative increase in purity)?
(c) Which of the purification procedures is least effective?
(d) Is there any indication based on the results shown in the table that the enzyme after step 6 is
now pure? What else could be done to estimate the purity of the enzyme preparation?
S-34 Chapter 3 Amino Acids, Peptides, and Proteins
Procedure Total protein (mg) Activity (units)
1. Crude extract 20,000 4,000,000
2. Precipitation (salt) 5,000 3,000,000
3. Precipitation (pH) 4,000 1,000,000
4. Ion-exchange chromatography 200 800,000
5. Affinity chromatography 50 750,000
6. Size-exclusion chromatography 45 675,000
2608T_ch03sm_S26-S43 2/1/08 11:45AM Page 34 ntt 102:WHQY028:Solutions Manual:Ch-03:

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