影响多肽与花粉钙调素亲和性的因素──多肽的圆二色性及核磁共振研究

钙调素 ( Ca M)存在于所有真核细胞生物体内 ,它可与很多天然的生物活性肽结合 ,如 β-内啡肽、胰高血糖素和某些昆虫毒液肽等 [1] .有关 Ca M与多肽相互作用的研究普遍认为 ,对 Ca M有高亲和性的多肽应该具有形成α螺旋结构的显著倾向[2 ] .为进一步确认多肽的主链构象对 Ca M亲和能力的影响 ,我们采用圆二色性光谱和核磁共振波谱分析了荞麦花粉碱性十二肽 BPP-1和它的类似物 BPP-3的结构特征 ,配合多肽对花粉钙调素 ( p Ca M)的结合能力 ,发现肽链的可塑性和 C端的极性是影响多肽与 p Ca M亲和能力的因素 ,而形成 α螺旋结构的倾…

The Factors Influencing the Peptide Affinity for Pollen Calmodulin Peptide Conformation Study by CD and 2D-NMR

Synthetic buckwheat pollen peptide BPP-1(A-P-V-L-Q-I-K-K-T-G-S-N)　and its analogues BPP-2[(D)A-P-V-L-Q-I-K-K-T-G-S-N-NH2], BPP-3(A-P-A-L-Q-L-K-K-N-G-S-Q-G-NH2)　showed different binding behavior to rape-pollen calmodulin(pCaM). Fluorescence titration experiments demonstrated that BPP-1 had no affinity for pCaM while its C-terminal amide, BPP-2, had fair affinity for pCaM(dissociation constant of pCaM-peptide complex, Kd=2.9×10－1　μmol/L) due to the decrease in C-terminal polarity. But compared with BPP-3(Kd=8.1×10－2　μmol/L), which was also a peptide amide, BPP-2 had a much lower binding ability. In order to investigate other factors influencing peptide affinity for pCaM besides polarity(or hydrophobicity), CD and 2D-NMR spectroscopes were used to study the conformations of BPP-1 and BPP-3. It revealed that molecular flexibility could affect peptides ability to bind pCaM. BPP-1 displayed rigid extended peptide bonds in the middle region where the basic amino acid pair Lys7-Lys8 is located, flanked by flexible peptide segments on both terminals; while the middle five-residue region of BPP-3 exhibited as very flexible segment. Such a structural character might facilitate BPP-3 to adopt a conformation contributive to the interaction of two Lys residues with the acidic residues of pCaM in its peptide-binding site and resulted in higher affinity. As this study revealed, both of the two peptides showed the lack of ordered structure in the aqueous solution. It seemed that there was no relationship between peptide affinity for CaM and the propensity of peptide chain to form α-helix. This contradicted what most previous researches approved that　α-helix was an important character of peptide with high affinity for CaM.