Identification of important amino acid residues of the Na+-Ca2+ exchanger inhibitory peptide, XIP.
Identification of important amino acid residues of the Na+-Ca2+ exchanger inhibitory peptide, XIP.
复制标题
Na -Ca2 交换抑制肽 XIP 的重要氨基酸残基的鉴定。
DOI：
10.1007/s002329900197
复制
发表时间：
1997
期刊：
影响因子：
--
通讯作者：
Philipson,KD
中科院分区：
文献类型：
--
作者：
He,Z;Petesch,N;Voges,K;Röben,W;Philipson,KD
Na+-Ca~(2+)交换器在兴奋-收缩偶联过程中通过跨膜转运Ca~(2+)而在心肌收缩能力中发挥重要作用。一个20个氨基酸的多肽，XIP，被合成来模拟交换区域，抑制交换活性。我们在这里确定了对抑制功能重要的氨基酸残基。测定了修饰多肽对Na~+-Ca~(2+)交换活性的影响。交换活性被评估为45Ca~(2+)对Na~+负载的心肌细胞膜小泡的摄取。我们发现，XIP的全长对于最大效力是重要的，尽管主要的抑制组分在5到16个残基之间。碱性残基和芳香族残基对XIP的抑制功能最重要。精氨酸12和精氨酸14被丙氨酸或谷氨酰胺取代，极大地降低了XIP的效力，表明这些残基在可能的电荷-电荷相互作用中发挥了关键作用。用丙氨酸或谷氨酸取代其他碱性残基对XIP的效力影响较小。所有芳香族残基都参与了与交换器的结合，可能是通过色氨酸荧光表明的疏水相互作用。最大抑制作用需要6位酪氨酸，苯丙氨酸5位和8位酪氨酸只能被其他芳香族残基取代。酪氨酸10和酪氨酸13可以被其他较大的残基取代。XIP的特定构象，以及分子所有部分提供的结构约束，是发挥最佳抑制功能所必需的。
The Na+-Ca2+exchanger plays an important role in cardiac contractility by moving Ca2+across the plasma membrane during excitation-contraction coupling. A 20 amino acid peptide, XIP, synthesized to mimic a region of the exchanger, inhibits exchange activity. We identify here amino acid residues important for inhibitory function. Effects of modified peptides on Na+-Ca2+exchange activity were determined. Exchange activity was assessed as45Ca2+uptake into Na+-loaded cardiac sarcolemmal vesicles. We find that the entire length of XIP is important for maximal potency, though the major inhibitory components are between residues 5 and 16. Basic and aromatic residues are most important for the inhibitory function of XIP. Substitutions of arginine 12 and arginine 14 with alanine or glutamine dramatically decrease the potency of XIP, suggesting that these residues play a key role in possible charge-charge interactions. Substitutions of other basic residues with alanines or glutamines had less effect on the potency of XIP. All aromatic residues participate in binding with the exchanger, probably via hydrophobic interactions as indicated by tryptophan fluorescence. A tyrosine is required at position 6 for maximal inhibition and phenylalanine 5 and tyrosine 8 can only be replaced by other aromatic residues. Tyrosine 10 and tyrosine 13 can be replaced with other bulky residues. A specific conformation of XIP, with structural constrains provided by all parts of the molecule, is required for optimal inhibitory function.