COMMUNICATIONS TO THE EDITOR
5028
1 13 H~N-lys-val-phe-gly-arg-cys-glu-leu-ala-ala-ala-met-lys
I
I
arg
PHZ ,NHz I try-asp-gly-leu-ser-tyr-gly-arg-tyr-asp-asp-leugly- h i s II
val
I/
33 ,NHz ,NHz ,NHz cys-ala-ala-lys-phe-glu-ser-asp-phe-asp-thr-glu-ala-thr
&
asp-thr-gly-lys
I
'
val
tYr
cys -
glu
1
116
I
I
r"z
I "rz asp rXHs I asp-thr-ser-gly-asp-thr-asp-arg
1
rNHz rNHz gly-ileu-leu-glu-ileu-asp-ser-arg
I
I
I
!
ala
r"zl
arg
I
gly-pro-thr-arg-gly-asp-asp-cys-try
I
I
'7
try
I
sr
arg-asp-leu-cys-asp-Ileu-pro-c?s-ser ,NHz 1 NH?
rkHz
I
ala-thr-ileu-asp-ser-ser -leu- leu - a l a arg gly
I
TI
arg
I
I
afg
ser
'r
Val
I 1
I
129 ala-Val-try-ala-asp-met-gly-asp-gly-asp leu-COOH Fig. 1.-Schematic representation of points of attachment of a,a'-dibromo-p-xylenesulfonicacid.
Further purification of the labeled ~teptideswas then followed by removal of the cross-linking residues under conditions used to hydrogenolyze benzyl amines. l 6 TABLE I QUALITATIVE ASALYS~S O F PEPTIDES INVOLVED LINKAGES Peptide
Cross linkage,a lysine residue
96
-
Ih'
CROSS
Aniino acids
97
-+
+
-+
(16) I,. Birkofer, B e y . d e u l . chem Ges., 7 6 , 429 (1942).
DEPARTMENT OF CHEMISTRY UNIVERSITY O F CINCINXATI CINCIXNATI 21, OHIO
c. B. HIREMATH RICHARD A. DAY
RECEIVED AUGUST21, 1964
Radical Scavenging in the Radiolysis of Liquid Methanol-Benzene Mixtures
Sir:
Calcd.: ala, arg, asp, gly, leu, met, ser, val Found: ala, arg, asp, gly, ileu,b met, ser, val Calcd.: lys AI2 96 9i Found: lys Calcd.. ala, asp, cys, ileu, leu, lys, Air 96 97 pro, ser, thr, try, Val Found: ala, asp, cys, ileu, leu, lys, pro, ser, thr, . ,' val Calcd.: ala, asp, cys, gly, leu, lys, Bi1 33 116 ser, try, tyr, val Found: ala, asp, cys, gly, leu, lys, .,' tyr, val ser, Calcd.: ala, arg, asp, glu, phe, ser, t h r Bi2 33 116 Found: ala, arg, asp, glu, phe, ser, thr Calcd. : cys, lys Bn 33 116 Found: cys, lys Calcd.: ala, arg, asp, glu, gly, ileu, B ~ z 33 + 116 thr, try, val Found: ala, arg, asp, glu, gly, ileu, t h r , . . . ,' val r\ccording to Canfield's sequence Analysis ambiguous with respect to differentiating between leucine and isoleucine. Acid hydrolysis left no detectable tryptophan residues. A11
Peptide A afforded only one nonradioactive peptide (AI) upon hydrogenolysis and peptide B gave two nonradioactive peptides ( B 1 and Bz). Peptide C was present in such small amounts that i t was neglected. Peptide A1 upon trypsin digestion yielded three new peptides (All, A12, and A13) the analysis of which (Table I) showed that they were derived from amino acids 7 3 to 112 in the primary sequence announced by CanfieldI7 (cf. Fig. 1). The three tryptides in this region correspond to Canfield's Tll, Tlz, and T13. Peptides B1 and BS upon tryptic digestion each led to two new peptides (Bll, Biz, B2', and B2*),respectively. These were analyzed and shown to correspond to Canfield's respectively (see Fig. 1 tryptides T6,T7,TlSrand TT6, and Table I ) . I t follows, then, t h a t DBX introduced cross-links into lysozyme and in particular between the €-amino groups of lysine residues 96-97 and X - l l ( j . The link between residues 3 3 and 116 corresponds to an assignment made by Herzig, et al., l 2 for phenal-2,1disulfonyl chloride, a reagent of nearly identical span. The possible significance in the difference in patterns of cross-linking by reagents of nearly identical dimensions of greatly different reactivities will be discussed elsewhere. (17) I
