(Bio Mathematical Lab of Sikander Aqeel)
CHAPTER 3 PROTEINS
Proteins
(2/26/2016)
Amino Acid Sequence of Polypeptide Chain:
The next step is to identify the amino-terminal and the carboxyl-terminal residues of the polypeptide chain, the first successful method, developed by F. Sanger utilized the reaction between the free a-amino group of the amino-terminal residue of the chain with 2.4 dinitrofluorobenzene to form a yellow 2,4 dinitrophenyl (DNP) derivative, when such a DNP derivative of a polypeptide is subjected to hydrolysis with 6 N HCl, all the peptide bonds are hydrolyzed, however the bond between 2,4 dinitrophenyl group,
Tetrapeptide = C18H10N4O5 = 216 + 12.5 + 56 + 80 = 364.5 mole
Hydrogen chloride = HCl = 36.25 mole
For example
= a2 + b2 = c2
= a2 = CHl and b2 = 24 hours = c2
= a2 = 1.25 (H) + 35 (Cl) and b2 = 24 = c2
= a2 = 36.25 + b2 = 24 = c2
= a36.25 + b24 = c2
= 1314.0625 + 576 = c2
= a + b = 1890.0625 = c2
= a + b = 1890.0625 / 43.5 = 43.5
= 43.5 * 8.37931 FDNB = 364.499985, (FDNB C5H10N2O4 = 164.5 / 18.5 water = 8.8918)
(FDNB) 364.5 mole = Tetrapeptide = HF
(FDNB) 364.5 mole – 1.25 = 363.25 (2,4 dinitrophenyl Tetrapeptide)
(FDNB) 363.25 mole --- 3H2O = 55.5 + 30.25 P -------- 85.75 m/g = C3H3NO2 = (2.4 dinitrophenyl-amino acid corresponding to the NH2 terminal residue)
2.4 dinitrophenyl-amino acid corresponding = 85.75 + 333 mole = 418.75 mole free amino acids,
Free amino acids = C15H12N3O6 = 180 + 15 + 42 + 96 = 333
Free amino acids = 333 mole
2.4 dinitrophenyl = 85.75
Total = 418.75 mole free amino acids
Total answer is
FDNB—HF— Tetrapeptide---- FDNB 2,4 dinitrophenyl Tetrapeptide------- NH2—terminal corresponding
And it occurs from 8.89 mole water of FDNP system, which make able to amino acid corresponding for free amino acids, and this long polypeptide chain, can be “sequenced” automatically by the Sikande Aqeel method, because when such a DNP derivative of a polypeptide is subjected to hydrolysis with 6 N HCl, all the peptide bonds are hydrolyzed, however the bond between 2,4 dinitrophenyl group,
(2/27/2016)
The a-amino group of the amino-terminal amino acid is relatively stable to acid hydrolysis, Consequently, the hydrolyzed will contain all the residues of the polypeptide chain as free amino acids except the amino-terminal amino acids, which will be present as 2,4-dinitrophenyl derivative, this can be easily separated from the unsubtituted amino acids and identified by chromatographic comparison with authentic DNP derivative of the different amino acids,
More recently! Greatly improved and more quantitative methods, for determination of the amino-terminal amino acid residue have been developed, in particular mouthed of P Sikander Aqeel, in which the terminal amino acid is converted into the phenylthiohydantoin derivative, the basis of this method,
P Sikander Aqeel
Tetrapeptide = C18H10N4O5 = 216 + 12.5 + 56 + 80 = 364.5 mole
Hydrogen chloride = HCl = 36.25 mole
For example
= a2 + b2 = c2
= a2 = HCl and b2 = 24 hours = c2
= a2 = 1.25 (H) + 35 (Cl) and b2 = 24 = c2
= a2 = 36.25 + b2 = 24 = c2
= a36.25 + b24 = c2
= 1314.0625 + 576 = c2
= a + b = 1890.0625 = c2
= a + b = 1890.0625 / 43.5 = 43.5
= 43.5 * 8.5 phenyliso thiocanate = 369.75,
= 369.75 - 5.25 = 364.5 mole, and (phenyliso thiocanate C8H12NS = 157.25 / 18.5 water = 8.5)
(phenyliso thiocanate) 364.5 mole = Tetrapeptide
(phenyliso thiocanate) 364.5 – 17.25 OH = 347.25 (phenylthiocarbamyl Tetrapeptide)
(phenylthiocarbamyl Tetrapeptide) 347.25 mole --- HCl = 36.25 + 30.25 P + 166.5 [9(H2O)]-------- 233 m/g - 5.5 = 227.5 mole Phenylthiohdantoin derivative of NH2-terminal amino acid,
Phenylthiohdantoin = C10H14N3O1S1 = 120 + 17.5 + 42 + 16 + 32 = 227.5
Phenylthiohdantoin = 227.5 * 1.6021 % NH2-terminal act = 364.47775
= 364.5 = phenyliso thiocanate = C18H10N4O5
Phenylthiohdantoin = 227.5
phenyliso thiocanate = 364.5 +
Total = 592 mole Original peptide minus NH2-terminal residue,
Total answer is
Which of main part is 166.5 mole water with P method of sikander Aqeel and Hydrogen chloride, So the Tetrapeptide go in phenylthiocarbamyl Tetrapeptide to less one OH ion, and phenylthiocarbamyl Tetrapeptide go in Original peptide to less HCl, for the carboxyl-terminal, for more easily understand, supposed you have 592 mole peptide into the sealed glass, now how many types of peptide are present into the sealed glass.
Sealed glass = 592 mole / 120 centigrade temperature = 4.93
= 5 peptide chains are present into the sealed glass
1 = phenyliso thiocanate
2 = Tetrapeptide
3 = phenylthiocarbamyl Tetrapeptide
4 = Phenylthiohdantoin
5 = Original peptide
Now you should never forget that two chromosomes are jointed with each another into the cell for make pair of metabolism, if two chromosomes are feel presences of Carboxyl group then the Carbon function will be activate into the one pair of chromosomes,
Carboxyl group = COOH = 45.25 * 12 activate carbon atom = 543
= 543 + 49 [3(OH ion)] = 592
= 592 mole Original peptide
This Sikander Aqeel method has used to identify the carboxyl-terminal residue, in one the terminal carboxyl group of the polypeptide is reduced to a hydroxyl group, I am writing first times this bio-chemistry, when I will write four times this bio-chemistry then your medical science would be too much easy to different treatments of human body, and it is very necessary to world health care,
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