TO THE
EDITOR
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A KEY pyridine/HOAc/HzO (30 :20 :6 :24) or n-BuOH/ 1-METHYL-6-ETHYL-3-AZAPHENANTHFU3NE9 DEGRADATION PRODUCT OF ATISINE HOAc/H20 (4:1:5) solvent systems. Each purified fraction was submitted to N-terminal residue anal- Si?: residue analysisj6 quantitay s i ~ t,o~ C-terminal ~ ~ The recent correlation of the aconitum alkaloid tive amino acid analysis,8 and finally to sequential atidine'J and the delphinium alkaloid ajaconine2m3 degradation from the N-terminus by the Edman with atisine emphasizes the key position this subm e t h ~ d . ~The . ~ following peptide fragments were stance occupies among the diterpene alkaloids of Lys.Arg.Arg.- the two genera. Recently structures I and I1 have identified: Gly.Lys.Pro.Val.Gly.Lys; Pro.Va1; Arg.Try; Ser.Met.Glu.His.Phe; Ser.- been suggested for d i h y d r ~ a t i s i n eand ~ atisine,6 Tyr ; Ala. Phe.Pro.Leu ; Lys (Val,Tyr,Pro,AspzGly,- mainly on the basis of the striking analogy of the Glu2,Ala).Ser.Ala.Glu.NH2 ; Lys. (Va1,Tyr, Pro,Xsp2,- chemistry of these substances7 to that of the garrya Gly,Glu&laa,Ser,Phe) and Glu.Phe. alkaloids8 Subsequent experimental work has Tryptic digests of the hormone (substrate/ demonstrated the presence of the oxazolidine enzyme = 90/1 (w./w.), p H 9-9.5, 40" for 6 hours) were submitted to countercurrent distribution for 37 transfers in the n-BuOH/20% HOAc system. The material with a partition coefficient ( K ) of 2 (peptide T2) was isolated and was shown to be homogeneous by N-terminal and amino acid analyses. Sequential degradation51g of this peptide moiety9,10,11 in atisine and isoatisine, the /hiniriofrom the N-terminus, together with analysis by the ethanol group11,12in dihydroatisine and the disdinitrophenylation method7J gave the structure : position of the D-ring and its substituents.13 An Val. Tyr. Pro. Asp (Gly, Glud, Alas, Asp, Ser, Phel, important piece of evidence bearing on the skeleton Pro, Leu). Partial acid hydrolysis (3 M HC1, 24 of atisine is the structure of the CI6Hl5Nbase14 (obhours a t 40") of this material yielded the following tained on selenium dehydrogenation) which conpep tides : Ala.Glu.ilsp; Gly .Glu (Alaz,Glu ,Asp,Ser) tains all but six of the carbon atoms of atisine and and (Val,Tyr,Pro,Asp, Gly,Glu).Ala. relates the heterocyclic ring to the rest of the moleThe remainder of the material from the counter cule. LlTe now wish to report the identification of current distribution of tryptic digests of bovine this base as l-methyl-6-ethyl-3-azaphenanthrene corticotropin was isolated and further separated (VII)'j by an unambiguous synthesis from 7 by zone electrophoresis and paper chromatography ethyltetralone-1 (111). This synthesis provides by means of the techniques employed for the the first evidence fixing the position of the nitrogen chymotryptic digests. The peptide fragments atom with respect to the rest of the atisine molelisted were identified: Ser.(Tyr,Ser,Met,Glu,His,-cule. Phe) .Arg ; Try.Gly .Lys.Pro.Va1.Gly .Lys ; L ys .Xrg Alkylation of the pyrrolidine enamine of ;-ethyland Arg.Arg.Pro.Va1.Lys. tetralone-1'6 (111) \Tith ethyl a-iodopropionate was From the above data, an amino acid sequence is effected by the method of Stork" to give after proposed for bovine corticotropin 11) S. \\'. Pelletier, C h e m i s t r y arid I n d u s t r y , 1016 1 1 0 i O ) . Ser.Tyr.Ser.Met.Glu.His.Plie..lrg.Try.~l~.L~s.Pr~.~~~ ~l. ( 2 ) S . \V.Pelletier, Science, 126, 1234 (1957); Cheinislry a r i d I n d i r s 1 2 3 4 5 6 7 8 9 1 0 1 1 1 2 1 3 I r y , lij70 (1957). ( 8 ) D. Dvornik and 0 . E . Edwards, Cnnnd. J . Chcrn., 36, SFO Gly.L\s.Lys..~rg..L\rg.Pro.T'al.Lys.Val.Tyr.Pro..L\s~).GI?. (19,573. 11 15 16 17 18 19 20 21 22 23 24 25 %ii (4) U. Dvornik and 0. E . Edwards, C h e m i s t r y a m i I n d u s t r y , 952 (1937). KH2
I
( 5 ) K Weisner, R. .4rmstrong, >I. F. Bartlett and J . A . Edwards, 1339 (1954). ?. FI . .Bartlett, P h . D . Thesis Summary, University of New Brunswick, SIay, 19.il. (7) For leading references see W..4. Jacobs, J . Org. Cherll., 16, 1593 ( 1!l5 1). ( 8 ) K . Weisner, el n l . , (a) C a i i n d . J . Chein., 30, (308 (19.52); (1)) B w . . 86, 800 (1953); (c) THIS J O U R N A L , 76, 0068 (1054); (cl) E x f i w i c i i t i a , 11, 255 (1955). ($1) S. W. Pelletier and TV, .2. Jacobs, THIS J O U R N A L 76, 4196 (195%). (10) S. \\I. Pelletier and \V. A . Jacobs, C1ieinzsli.y aiad Iizdiistry, 1385 (1955). (11) 0 . E. Edwards and T Singh, Cairad. J . Chern., 33, 418 ( I B S ) , ibid., 32, 405 (1954). ( 1 2 ) S. W. Pelletier and W. A. Jacobs, THIS J O U R N A L , 78, 41-11 (1956). (13) S . W.Pelletier and W. A . Jacobs, i b i d . , 78, 413R (19.X). ( 1 4 ) W. A . Jacobs and L. C. Craig, J . R i d . C h e n . . 143, 589 ( 1 9 l 2 ) . (15,) This structure was suggested b y the similarity of the ultra\i
