Sanger

Yellow liquid m.p. 26" C, b.p. l37 "C/2mm. Reagent introduced by Sanger for identification of the terminal amino group of peptides with which it forms an /V-2,4-dlni-irophenyl derivative under mild conditions. 

The compound 1 fluoro 2 4 dinitrobenzene is exceed ingly reactive toward nucleophilic aromatic substi tution and was used in an imaginative way by Frederick Sanger (Section 27 11) in his determination of the struc ture of insulin... 

Sanger was a corecipient of a second Nobel Prize in 1980 for devising methods for sequencing nucleic acids Sanger s strategy for nucleic acid sequencing will be described in Section 28 14... 

Several chemical methods have been devised for identifying the N terminal ammo acid They all take advantage of the fact that the N terminal ammo group is free and can act as a nucleophile The a ammo groups of all the other ammo acids are part of amide linkages are not free and are much less nucleophilic Sanger s method for N terminal residue analysis involves treating a peptide with 1 fluoro 2 4 dimtrobenzene which is very reactive toward nucleophilic aromatic substitution (Chapter 23)... 

Fluoro 2 4 dimtrobenzene IS commonly referred to as Sanger s reagent... 

Sanger also determined the sequence of the A chain and identified the cysteine residues involved m disulfide bonds befween fhe A and B chains as well as m fhe disulfide linkage wifhin fhe A chain The complefe insulin sfruefure is shown m Figure 27 11 The sfruefure shown is fhaf of bovine insulin (from cattle) The A chains of human insulin and bovine insulin differ m only fwo ammo acid residues fheir B chains are identical except for the ammo acid at the C terminus... 

The years that have passed since Sanger determined the structure of insulin have seen refinements m technique while retaining the same overall strategy Enzyme cat alyzed hydrolysis to convert a large peptide to smaller fragments remains an important... 

When Sanger s method for N terminal residue analysis was discussed you may have wondered why it was not done sequentially Simply start at the N terminus and work steadily back to the C terminus identifying one ammo acid after another The idea is fine but It just doesn t work well m practice at least with 1 fluoro 2 4 dimtrobenzene... 

Carboxypeptidase catalyzed hydrolysis can be used to identify the C terminal ammo acid The N terminus is determined by chemical means One reagent used for this purpose is Sanger s reagent 1 fluoro 2 4 dimtrobenzene (see Figure 27 9)... 

FIG. 14-109 Typical separators using impingement in addition to centrifugal force, a) Hi-eF purifier. (V D, Anderson Co.) (h) Flick separator. (Wutster line separator. (Centrifix Co7j>., Bull. 220.)... 

Dinitrofluorobenzene (Sanger s reagent) [70-34-8] M 186.1, m 25-27 , b 133 /2mm, 140-141 /5mm, d 1.483. Crystd from ether or EtOH. Vacuum distd through a Todd Column (see p. 174). If it is to be purified by distn in vacuo, the distn unit must be allowed to cool before air is allowed into the apparatus otherwise the residue carbonises spontaneously and an EXPLOSION may occur. The material is a skin irritant and may cause serious dermatitis. 

As the molecular structure of DNA was being elucidated, scientists made significant contributions to revealing the structures of proteins and enzymes. Sanger [2] resolved the... 

One of the historically most significant examples of aromatie nueleophilie substitution is the reaetion of amines with 2,4-dinitrofluorobenzene. This reaetion was used by Sanger to develop a method for identifieation of the N-terminal amino acid in proteins and the proeess opened the way for struetural eharacterization of proteins and other biopolymers. 

Acid fluondes such as oxalyl fluoride, tnchloroacetyl fluonde, propionyl fluonde, as well as Sanger s reagent, 2,4-dinitrofluorobenzene, easily cleave N,N-acetals (aminals) to give high yields of fluoromethyldialkylamines [82] (equation 20) (Table 5)... 

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