214 C. H. W. HIRS, WILLIAM H. STEIN & STANFORD MOORE [12 

 linked, detailed structural studies of all of these peptides were not required. 

 It was possible to select for sequence work only those peptides of suitable 

 size which do not overlap one another (Fig. 1). The starting point was the 

 series of 13 peptides obtained by tryptic hydrolysis of oxidized ribonuclease. 

 These peptides account for all of the ribonuclease molecule, and can be 



Lys.Glu.Thr.Ala.Ala.Ala.Lys.Phf.Glu.Arg.Ser.Thi-.Ser.Ser.Asp.His.Met.Glu.Ala.Ala. 



© 



rNH2 I rNH2-NH2 i-NHg 

 Ser.Ser.Asp.Ser.Tyr.Cys.Asp.Glu.Met.Met.Lys.Ser.Arg.Asp.Leu.Thr.Lys.Asp.Arg. 



© @ 



I rNH2 r-NH2 rNH2^ i pNH, 

 (Cys.Asp,Val,Pro,Thr,Lys).Phe.(Glu,Val,Leu.Ser,His).(Asp,Glu,Ala,Val).(Cys,Glu,Ala,Val,Ser). 



® " ? 



pNH2 I rNH2 r-NH2-NH2 rNH2 



I I I I I I I 



Lys. Asp. Val. Ala. Cys. Lys. Asp. Gly.Thr. Asp. Glu. Cys.Tyr. Glu. Ser.Tyr.Ser.Thr. Met. Ser. lieu. Thr. 



y [-NH2 I pNH2-NH2 

 Asp.Cys.Arg.Glu.Ser. Thr. Ser. Gly. Lys. Tyr. Pro. Asp. Ala. Cys.Tyr. Lys. Thr. Thr. Asp. Glu. Ala. Lys. 



@ 



I n 



(Val.Ileu2,His).(Cys.Asp,Glu.Gly.Ala,Pro).Tyr.(Val2.Pro,His). Phe.Asp.Ala.Ser.Val 



Fig. 2. Arrangement of the amino acid residues in oxidized ribonuclease.^ The sequences 

 determined to date are underlined. The residues in parentheses are of undetermined order. 

 The cysteic acid residues have been numbered consecutively, I to VIII, starting from the 

 amino-end of the chain. 



prepared in yields ranging from 50 to 100 per cent. Nine of these 13 pep- 

 tides, varying in size from dipeptides to heptapeptides, account for 42 of 

 the 124 residues in the chain, and are small enough to be studied directly. 

 The structures of all nine have been worked out. The remaining four pep- 

 tides of the trypsin series, O-Tryp 4, 0-Tryp 9, O-Tryp 2, and 0-Tryp 16 

 (Fig. 1), which contain 21, 22, 19, and 20 residues respectively, account 

 for the rest of the molecule, but are too large to be useful, as such, for 

 sequence studies. However, from the knowledge of the points of cleavage 

 by chymotrypsin provided by the partial structural formula, it can be pre- 

 dicted that each of these four large peptides should be split individually 

 with chymotrypsin into fragments of a more manageable size. This has been 

 carried out, and in each instance the expected segments were obtained. For 

 example, the action of chymotrypsin on O-Tryp 4, which contains 21 amino 



