Matuszek M A, Comis A, Burcher E
School of Physiology and Pharmacology, University of New South Wales, Sydney, Australia.
Pharmacology. 1999 May;58(5):227-35. doi: 10.1159/000028285.
Structure-activity relationships of neurokinin A (NKA) and the two analogues NKA(4-10) and [Nle10]NKA(4-10) were investigated at the rat fundus NK-2 receptor, using selected amino acid substitutions. Both radioligand binding with [125I][Lys5,Tyr(I2)7,MeLeu9, Nle10] NKA(4-10) and functional studies were performed and correlated. In membrane binding experiments loss of His1 and Lys2, or replacement of Lys2 with Ala did not substantially alter binding affinity of NKA. NKA(4-10) free acid was unable to compete with the radioligand. [Nle10]NKA(4-10) binding affinity to rat fundus membrane preparations was decreased when substituting Asp4 with Gln or Asn, or Val7 with either Tyr or Ile. Replacement of Ser5 with the negatively charged Glu also decreased the binding affinity, but substitution with the positively charged Lys substantially increased the affinity of [Nle10] NKA(4-10) for the NK-2 receptor. Lengthening NKA(4-10) or [Nle10]NKA(4-10) with Ala11 or Nle11, respectively, decreased the binding affinity of the peptide. In both binding and functional studies, replacement of any of the residues of NKA(4-10), except for Ser5, with alanine decreased the affinity of the peptide for the NK-2 receptor. Ala substitutions at positions 4, 6, and very obviously at 8, 9 and 10 of NKA(4-10) yielded peptides unable to achieve a maximum contractile response, although they did not demonstrate antagonist activity. These data confirm the importance of the NKA carboxyl terminus, and the requirement for Phe6, Val7, Gly8, Leu9 and Met10 integrity for interaction with the NK-2 receptor. They also suggest that Ser5 is a good site to target modifications leading to the design of new potential drugs.
利用选定的氨基酸取代，在大鼠胃底NK-2受体上研究了神经激肽A（NKA）以及两种类似物NKA(4-10)和[Nle10]NKA(4-10)的构效关系。进行了放射性配体与[125I][Lys5,Tyr(I2)7,MeLeu9,Nle10]NKA(4-10)的结合以及功能研究，并将二者关联起来。在膜结合实验中，His1和Lys2缺失，或者Lys2被Ala取代，均未显著改变NKA的结合亲和力。NKA(4-10)游离酸无法与放射性配体竞争。当用Gln或Asn取代Asp4，或者用Tyr或Ile取代Val7时，[Nle10]NKA(4-10)与大鼠胃底膜制剂的结合亲和力降低。用带负电荷的Glu取代Ser5也降低了结合亲和力，但用带正电荷的Lys取代则显著增加了[Nle10]NKA(4-10)对NK-2受体的亲和力。分别用Ala11或Nle11延长NKA(4-10)或[Nle10]NKA(4-10)会降低该肽的结合亲和力。在结合和功能研究中，除Ser5外，用丙氨酸取代NKA(4-10)的任何残基都会降低该肽对NK-2受体的亲和力。在NKA(4-10)的第4、6位，以及非常明显地在第8、9和10位用Ala取代得到的肽，尽管未表现出拮抗活性，但无法产生最大收缩反应。这些数据证实了NKA羧基末端的重要性，以及Phe6、Val7、Gly8、Leu9和Met10完整对于与NK-2受体相互作用的必要性。它们还表明Ser5是进行修饰以设计新的潜在药物的良好靶点。
