Tor f r on Vol . 29, No . 4/ 5, pp. 521- 526, 1991 . ~ 0041- 0101/ 91 53. 00+ . 00 Pr i nt ed in Gr at Bri t ai n. ® 1991 Per g~r oon Pr ep pk AMI NOACI D SEQUENCE OF A SNAKE VENOM TOXI N THAT BI NDS TO THE MUSCARI NI C ACETYLCHOLI NE RECEPTOR EVERT KARLSSON, 1 CARL RI SI NGER, 1 MI I CAEL J OLKKONEN, 1 CHRI STER WERNSTEDT Z and As nu ADEM 3 ' Depart ment of Bi ochem i st r y, Bi omedi cal Cent r e, Box 576, 751 23 Uppsal a, Sweden, ~Ludwi g I nst i t ut e f or Cancer Resear ch, Bi omedi cal Cent r e, Box 595, 75I 23 Uppsal a, Sweden, and sI nst i t ut e of Geri at ri c Medi ci ne, Kar ol i nsl ca I nst i t ut et , Huddi nge Hospi t al , 141 86 Huddi nge, Sweden ( Recei ved 22 l une 1990; accept t d 29 Oct ober 1990) E . KARLS . SON, C . RI SI NGER, M . J OLKKONEN, C . WERNSTEDT and A . ADEM . Am i no aci d sequence of a snake venom t oxi n t hat bi nds to t he muscari ni c acet yl chol i ne r ecept or . Toxi con 29, 521- 526, 1991 . - The gr een mamba, Dendr oaspi s angust i ceps, has t wo pr ot ei n t oxi ns t hat bi nd to t he muscari ni c acet yl chol i ne r ecept or . The sequence of muscari ni c t oxi n 2 was det er m i ned with an aut omat i c gas phase sequencer . The C- t er m i nal r esi due is As p as det er m i ned by hydr azi nol ysi s and am i no aci d anal ysi s . Toxi n 2 has 65 am i no aci d r esi dues and a f ormul a wei ght of 7040 . It is homol ogous to a l ar ge number of ot her snake venom t oxi ns as shor t a- neur ot oxi ns, car di ot oxi ns/ cyt ot oxi ns and angust i ceps- t ype t oxi ns of mamba venoms . The sequence is conf i rmed in t he accompanyi ng ar t i cl e ( DUCANCEL, F., RownN, E . G ., CASSART, E ., HARVEY, A . L ., MENEZ, A . and BOULAI N, J .-C. Toxi con 29, Slfi~ 520, 1991) . TARE ar e at l east t hr ee phar macol ogi cal ( M1, M2, M3) ( DODDS et al ., 1987 ; LEVI NE et al ., 1987 ; GOYAL, 1988) and f i ve st r uct ur al subt ypes (m l, . . . m5) ( BONNER et al ., 1987 ; PERALTA et al ., 1987 ; Luo et al ., 1989; GOYAL, 1989) of muscari ni c acet yl chol i ne r ecept or s . The m l subcl ass cor r esponds pr obabl y to M1, m2 to M2 and m3 to M3 . Subt ype M1 dom i nat es in br ai n, M2 in smoot h muscl e and M3 in car di ac muscl e (LEVI NE and BI RDSALL, 1989) . No l i gand is known t hat bi nds onl y t o one subcl ass of r ecept or s, but s ome synt het i c compounds ar e r at her sel ect i ve, e .g. pi r enzepi ne has a 10- 15 f ol d hi gher af l ' I ni t y f or M1 t han f or any ot her subt ype ( AKI BA et al ., 1988; BUCKLEY et al ., 1989 ; BONNER, 1989) . Muscari ni c t oxi ns f r om mamba venoms ar e t he onl y pr ot ei n t oxi ns known t hat bi nd t o t he muscari ni c acet yl chol i ne r ecept or . They i nhi bi t t he bi ndi ng of qui nucl i - di nyl benzi l at e ( QNB) to r at br ai n synapt osomes to about 50%( ADEMet al ., 1988) . QNB is an ant agoni st t hat bi nds to al l subt ypes of muscari ni c r ecept or s and t he par t i al i nhi bi t i on of t he bi ndi ng m i ght i ndi cat e t hat t he mamba t oxi ns ar e sel ect i ve f or one subt ype . Two muscari ni c t oxi ns wer e i sol at ed f r om Dendr oaspi s angust i ceps ( gr een mamba) venom ( pur chased f r om Jabr i a, B .V., Har der wi j k, The Net her l ands) as descr i bed ear l i er ( ADEM et al ., 1988) . The sequence of t oxi n 2, t he mai n t oxi n, was det er m i ned . The r educed and S- car boxymet hyl at ed ( RCM) der i vat i ve, pr epar ed accor di ng t o t he met hod Of CRESTF<ELD et al . ( 1963) , was di ssol ved in 0 .1 M ammoni umbi car bonat e ( pH 7 . 9) and 521