Determining the Amino Acid Sequence of Insulin Figure 3-24...

Determining the Amino Acid Sequence of Insulin Figure $3-24$ shows the amino acid sequence of bovine insulin. This structure was determined by Frederick Sanger and his coworkers. Most of this work is described in a series of articles published in the Biochemical Journal from 1945 to 1955. When Sanger and colleagues began their work in $1945,$ it was known that insulin was a small protein consisting of two or four polypeptide chains linked by disulfide bonds. Sanger's team had developed a few simple methods for studying protein sequences. Treatment with $F D N B$. FDNB (1-fluoro-2,4-dinitrobenzene) reacted with free amino (but not amide or guanidinium) groups in proteins to produce dinitrophenyl (DNP) derivatives of amino acids: (a) Explain how Sanger's results support his conclusions. (b) Are the results consistent with the known structure of 24)$?$

Determining the Amino Acid Sequence of Insulin Figure $3-24$ shows the amino acid sequence of bovine insulin. This structure was determined by Frederick Sanger and his coworkers. Most of this work is described in a series of articles published in the Biochemical Journal from 1945 to 1955.
When Sanger and colleagues began their work in $1945,$ it was known that insulin was a small protein consisting of two or four polypeptide chains linked by disulfide bonds. Sanger's team had developed a few simple methods for studying protein sequences.
Treatment with $F D N B$. FDNB (1-fluoro-2,4-dinitrobenzene) reacted with free amino (but not amide or guanidinium) groups in proteins to produce dinitrophenyl (DNP) derivatives of amino acids:
(a) Explain how Sanger's results support his conclusions.
(b) Are the results consistent with the known structure of 24)$?$

Key Concepts

Protein Sequencing Techniques

This concept involves methods used to determine the order of amino acids in a protein. In Sanger's work, this includes sequentially identifying the N-terminal residues of protein fragments after chemical modification. Such techniques allowed early researchers to piece together the complete amino acid sequence of proteins, an essential step in understanding protein structure and function.

Protein Structure Analysis

Understanding the overall structure of proteins, such as insulin, is key to determining how their amino acid sequence dictates their three-dimensional shape and biological activity. In the context of insulin, characterizing the different polypeptide chains and their connections was critical to establishing its functional form and the relationship between structure and function.

Disulfide Bond Formation

Disulfide bonds are covalent bonds that form between the sulfur atoms of cysteine residues in proteins, contributing significantly to the stabilization of the protein’s tertiary and quaternary structures. In insulin, these bonds link separate polypeptide chains, and recognizing their importance was crucial for Sanger's determination of the protein's connectivity and overall structure.

Chemical Derivatization with FDNB

FDNB (1-fluoro-2,4-dinitrobenzene) is a reagent used to chemically modify free amino groups in proteins, leading to the formation of dinitrophenyl (DNP) amino acid derivatives. This derivatization makes it possible to detect and identify amino acids sequentially, as only free amino groups react with FDNB. It was a fundamental step in Sanger's method for gradually determining the peptide sequence.

Transcript

00:02 In this question, they give us amino acid sequences of the foxp2 proteins for different organisms.

00:11 And these are listed here.

00:14 So first they ask us to highlight amino acids that are different.

00:18 Just by looking at them, we can see that number six has this amino acid that's different.

00:25 Number two has this one.

00:28 Number four has this one right here.

00:31 And then again, number four has this one right here.

00:35 So for letter a, we need to find which of these sequences are the same.

00:40 And these would be assigned to the chimpanzee, which is going to be labeled c, gorilla, which is going to be labeled g, and rhesus macaque, which is going to be labeled r.

00:52 And again, just by looking at them, we can see that number one, three, and five are the same.

00:59 So i'm going to go ahead and label them.

01:01 So c for chimpanzee, g for gorilla, and r.

01:06 For reese macoc.

01:08 For letter b, we need to find which of these is the human sequence, which is going to be labeled h.

01:15 And it's different at two amino acids.

01:18 So just by looking at them and based on the first step that we did, we can see that number four has two differences in amino acids.

01:26 So we're going to go ahead and label this one, number four.

01:30 We're going to label it human, h for human.

01:34 For letter c, we need to find the orangutan sequence, which is different from the cgo.

01:38 Are at one amino acid and it's different from the h at three amino acids.

01:44 So again, just by looking at it and based on the first step that we did, we can see that number six is different from the first, third, and fifth sequence at one amino acid...

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