269. 21949 330% [h] a1 ( i . M. Clorc. A M Gronenhorn. .I .M~i,!yi. R c i i i i 1982. 48. 402 -417: h) G. M. Chi-e. A. M GI-onenhorn. ihid 1983. 53. 423 ~442: c ) F. Xi, .I , M q y Rcwrio~i. 1992. !Zi. 651 656 [7] €5. Ernst, N. Cooke. P M. ,&her?. unpublished. The qrithetic tetrasaccharide corresponds to thc formula shown in the text.181 The cctndnm;iins of h u m a n E-selectiii were combined b! means of the poly-mera\echnin iretictimi (FCR) uith the Fc region of liuman I g G l , and subcloned iii the expression vector pcDNAl iieo (Invitrogen). Following trmsfection of CHO-KI cells (ATCC CCL-61) and selection with '3418 (Gibco). :i stahle cell line w s obtained which secreted c' r 25 pgni1. ' recombinant E-selectin IiIgG. For thc productimi of larger aniouiit~ t h i s cell line was cultivated in 3 hollo~r fiber hioreactor (1.1 in2. 50 mL, Cellco) in Opli. MEM culture medium (Ciihco) rupplementcd with 2 % fetal calf \ei-um nnd 200 m L -' gentamyciii. The supernatant o l~o n d i t i o n~d cclh was lir5t purified by d h t y chromatography on proteiii A agarme (Sigma). The proteiii fraction obtained hy elution with 20mM glqcine-HCl. pH 3.0. w a s iicutralired and chi-oin;itographed direct11 on an .inti-human-E-selectiii affinity column ( 3 mg monoclonal antibody 7A9 per mL affigel). Specifically horiiid E-selectin l g G was eluted nitli 5 v urea in buffered s a l i solution and finally dialyzed against Dubeccos PBS solution (PBS = phosphate buffered \aline). The protein thus obtained showed. i n SDS (sodium dccyl sull'atc) polyacry1;rmidc pel electrophoresis tinder reducing coiiditions. : i single band :it ca 140 kD. For NMR triialyris ca 6 me E-selectin hJgG wab diiilyxd against 50mM pcrdeutei-ated imidazol (Signs). I mh4 Ca-CI,. pH 7.4. iii D1O and concentrated by-means of Centricon YM-50 (Amicon) t o il Sinal volume ofO.5 mL. [O] a ) All NMR experiment.; w r e performed i n the Jnstitut fur Binpliqsikalisclie . . C;hemic (lei-Uniccrsitiit Fi-ankfiirt on :i Rruker DMXhOO specti-ometei-. We ~i s h to tliaiik Prof. Dr. H. Ruterjans for the the opportunilq to make the measurements. b) For the N MR eyperiments on the complex an E-selectin(1gG c1iiiner:i)'retrasacchnride ratio of 1: 15 M R S chosen. The concentration ofsialyl Letcis" \\>is 0.X1 inhi. that of E-selectin(lgG chimera) 5 4~~. [D,]Imidarolc (30iiihr) mas used as buffer. In addition the tolution contained NaCl (50mxi) and CaCI, (1 nib<). The pH w a s 7.4. Spinlock filtered NOESY spectra [lo] with mixing times T,,, of 25. 50. 75. 100. 125. 150. 175. and 200 ms were recorded at 600 MMr and 310 K. The relaxation lime way 1 X5 s. The spectral \bid111 was i000 H r (5 ppiii) in all casea. 512 Increments in T , Rere recorded. For each iiicrcment 32 transients were xcqnired each with 2K data points. After Lero-filling and multiplicirti~~n with squared coc functions in r l and T?. ii 2K x 1 K data matrix bas obtaincd by 2D Fourier transformation. T. Scherf. J Anglister. Biophr.~. .I 1993. 64. 7.54-761 NOESY specti-a o f lkcc F ciin(JgC~ chimera) and sialyl Lewis" tetr...
The catalytic CC coupling of chloroarenes by both the Heck and the Suzuki reactions has been achieved for the first time with new palladacycles of type 1, in which R can be, for instance, o‐tolyl or mesityl. They are an order of magnitude more active than conventional catalysts and, moreover, thermally considerably more robust. But above all, the cleavage of the PC bond and the deposition of palladium is not observed. Analogous reactions with bromoarenes can also be catalyzed to advantage with 1. The Heck reaction of haloarenes with vinyl derivatives (e.g., n‐butyl acryiate) gives (E)‐styrene derivatives in one step. In the Suzuki reaction the haloarenes are converted into biphenyl derivatives with phenylbornic acid, also in one step.
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