Proteinase-activated receptors: structural requirements for activity, receptor cross-reactivity, and receptor selectivity of receptor-activating peptides.
Proteinase-activated receptors: structural requirements for activity, receptor cross-reactivity, and receptor selectivity of receptor-activating peptides.
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蛋白酶激活受体：受体激活肽的活性、受体交叉反应性和受体选择性的结构要求。
DOI：
10.1139/y97-110
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发表时间：
1997
影响因子：
2.1
通讯作者：
Atsufumi Kawabata
中科院分区：
文献类型：
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作者：
M. Hollenberg;M. Saifeddine;B. Al‐Ani;Atsufumi Kawabata
我们已经使用了三种不同的生物测定系统（大鼠主动脉（RA）松弛;大鼠胃纵肌（LM）收缩;人胚肾293（HEK 293）细胞钙信号）来评估来自凝血酶受体（TRAP，基于人受体序列，SFLLRNPNDK.）或蛋白酶激活受体2（PAR 2AP，基于大鼠受体序列SLIGRL.）。我们的主要焦点是PAR 2 AP对PAR 2的激活和TRAPs对PAR 2的交叉激活。在RA和LM测定系统中，作为激动剂或拮抗剂，N-乙酰化（N-乙酰基-SLIGRL-NH 2）或具有反向N-末端序列（LSIGRL-NH 2）的PAR 2AP是无活性的。PAR 2AP的3位丙氨酸取代（SLAGRL-NH 2）导致生物活性的显著降低，1位苏氨酸取代丝氨酸（TLIGRL-NH 2）也是如此。然而，PAR 2AP位置4处的丙氨酸取代仅引起活性的适度降低，产生具有与人PAR 2AP SLIGKV-NH 2的效力相当的效力的肽（SLIARL-NH 2）。PAR 2AP在RA、LM和HEK测定系统中的效力顺序为SLIGRL-NH 2> SLIARL-NH 2> SLIGKV-NH 2> TLIGRL-NH 2> SLAGRL-NH 2。在HEK细胞中，没有PAR 2AP激活凝血酶受体（PAR 1）。然而，在HEK细胞试验中，TRAP，SFLLR-NH 2，激活或脱敏PAR 1和PAR 2受体，而非洲爪蟾TRAP，TFRIFD-NH 2，激活或脱敏选择性PAR 1，但不PAR 2。通过构建人-爪蟾杂合肽，我们发现TRAPs、TFLLR-NH 2和SFLLFD-NH 2选择性地激活HEK细胞中的凝血酶受体，而不激活或脱敏PAR 2。相反，TRAPs SFLLRD-NH 2和AFLLR-NH 2激活或脱敏PAR 1和PAR 2。在所有生物测定系统中TRAP的效力顺序为SFLLR-NH 2约等于SFLLRD-NH 2约等于TFLLR-NH 2> SFLLFD-NH 2> TFRIFD-NH 2。我们的结论是PAR 2AP的N端结构域以及位置3在PAR 2激活中起重要作用。相反，TRAP基序中的第一个和第五个氨基酸SFLLR-NH 2在激活凝血酶受体中不起独特作用，但如果适当修饰，则可以消除该肽交叉脱敏或激活PAR 2的能力，从而对PAR 1具有选择性。我们合成的PAR 1和PAR 2选择性肽将用于评估PAR 1和PAR 2受体系统在体内的作用。
We have used three distinct bioassay systems (rat aorta (RA) relaxation; rat gastric longitudinal muscle (LM) contraction; human embryonic kidney 293 (HEK293) cell calcium signal) to evaluate the activity and receptor selectivity of analogues of the receptor-activating peptides derived either from the thrombin receptor (TRAPs, based on the human receptor sequence, SFLLRNPNDK...) or the proteinase-activated receptor 2 (PAR2APs, based on the rat receptor sequence SLIGRL...). Our main focus was on the activation of PAR2 by PAR2APs and the cross-activation of PAR2 by the TRAPs. In the RA and LM assay systems, PAR2APs that were either N-acetylated (N-acetyl-SLIGRL-NH2) or had a reverse N-terminal sequence (LSIGRL-NH2) were inactive, either as agonists or antagonists. An alanine substitution at position 3 of the PAR2AP (SLAGRL-NH2) led to a dramatic reduction of biological activity, as did substitution of threonine for serine at position 1 (TLIGRL-NH2). However, alanine substitution at PAR2AP position 4 caused only a modest reduction in activity, resulting in a peptide (SLIARL-NH2) with a potency equivalent to that of the human PAR2AP, SLIGKV-NH2. The order of potency of the PAR2APs in the RA, LM, and HEK assay systems was SLIGRL-NH2 > SLIARL-NH2 > SLIGKV-NH2 > TLIGRL-NH2 > SLAGRL-NH2. In HEK cells, none of the PAR2APs activated the thrombin receptor (PAR1). However, in the HEK cell assay, the TRAP, SFLLR-NH2, activated or desensitized both PAR1 and PAR2 receptors, whereas the xenopus TRAP, TFRIFD-NH2, activated or desensitized selectively PAR1 but not PAR2. By constructing human-xenopus hybrid peptides, we found that the TRAPs, TFLLR-NH2, and SFLLFD-NH2 selectively activated the thrombin receptor in HEK cells without activating or desensitizing PAR2. In contrast, the TRAPs SFLLRD-NH2 and AFLLR-NH2 activated or desensitized both PAR1 and PAR2. The order of potency for the TRAPs in all bioassay systems was SFLLR-NH2 approximately equal to SFLLRD-NH2 approximately equal to TFLLR-NH2 > SFLLFD-NH2 > TFRIFD-NH2. We conclude that the N-terminal domain of the PAR2AP as well as positon 3 plays important roles for PAR2 activation. In contrast, the first and fifth amino acids in the TRAP motif, SFLLR-NH2, do not play a unique role in activating the thrombin receptor, but if appropriately modified can abrogate the ability of this peptide to cross-desensitize or activate PAR2, so as to be selective for PAR1. The PAR1- and PAR2-selective peptides that we have synthesized will be of use for the evaluation of the roles of the PAR1 and PAR2 receptor systems in vivo.
受体衍生肽激活血小板凝血酶受体的结构-功能关系。
DOI：
--
发表时间：
1992
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
VassalloJr,RR;Kieber-Emmons,T;Cichowski,K;Brass,LF
通讯作者：
Brass,LF