Esterolytic peptide dendrimers with a hydrophobic core and catalytic residues at the surface
Esterolytic peptide dendrimers with a hydrophobic core and catalytic residues at the surface
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DOI：
10.1002/adsc.200404070
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发表时间：
2004-08-01
影响因子：
5.4
通讯作者：
Reymond, JL
中科院分区：
文献类型：
--
作者：
Clouet, A;Darbre, T;Reymond, JL
以（R）-3-氨基-1，2-丙二醇为原料，合成了（3S）-4-（9-芴甲氧羰基氨基）-3-甲基（烯丙氧羰基）氨基乙氧基乙酸（1），并以此为支化单元合成了具有6个独立可变氨基酸位置的第二代肽树枝状聚合物。制备了三对非对映体树枝状聚合物，其具有共同的疏水核心和表面处的催化三联体氨基酸天冬氨酸、组氨酸和丝氨酸的排列。在pH6.0的缓冲溶液中，表面带有两个组氨酸残基的树枝状大分子催化荧光8-酰氧基-1，3，6-三磺酸芘酯的水解，其速率增强系数k（cat）/k（uncat）在10（3）范围内，米氏常数K-m在10（-4）M范围内。底物识别涉及静电相互作用，如观察到的芘-1，3，6，8-四磺酸盐的催化竞争性抑制所示。最活跃的树枝状聚合物中丁酰基酯和壬酰基酯之间的Km降低4倍至7倍，这为底物结合中的疏水组分提供了证据，这在密切相关的、活性较低的非对映体肽树枝状聚合物中不存在。
(3S)-4-(9-Fluorenylmethoxycarbonylamino)-3-methyl(allyloxycarbonyl)aminoethyloxyacetic acid (1) was prepared from (R)-3-anainopropane-1,2-diol and used as branching unit for the synthesis of second generation peptide dendrimers with six individually addressable variable amino acid positions. Three pairs of diastereomeric dendrimers were prepared bearing a common hydrophobic core and permutations of the catalytic triad amino acids aspartate, histidine and serine at the surface. Dendrimers with two surface histidine residues catalyzed the hydrolysis of fluorogenic 8-acyloxypyrene-1,3,6-trisulfonates in aqueous buffer pH 6.0 with rate enhancement k(cat)/k(uncat) in the 10(3) range and Michaelis-Menten constants K-m in the 10(-4) M range. Substrate recognition involves electrostatic interactions, as shown by competitive inhibition of catalysis observed with pyrene-1,3,6,8-tetrasulfonate. The 4-fold to 7-fold lowering in Km between the butyryl and nonanoyl esters in the most active dendrimers provides evidence for a hydrophobic component in substrate binding, which is absent in a closely related, less active diastereomeric peptide dendrimer.