β-Lactoglobulin variants as potential carriers of pramoxine: Comprehensive structural and biophysical studies

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Journal of Molecular Recognition Pub Date : 2023-08-23 DOI:10.1002/jmr.3052

Piotr Bonarek, Dorota Mularczyk, Joanna I. Loch, Katarzyna Kurpiewska, Marta Dziedzicka-Wasylewska

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引用次数: 0

Abstract

β-Lactoglobulin (BLG) is a member of the lipocalin family. As other proteins from this group, BLG can be modified to bind specifically compounds of medical interests. The aim of this study was to evaluate the role of two mutations, L39Y and L58F, in the binding of topical anesthetic pramoxine (PRM) to β-lactoglobulin. Circular dichroism spectroscopy, isothermal titration calorimetry (ITC), and X-ray crystallography were used to understand the mechanisms of BLG–PRM interactions. Studies were performed for three new BLG mutants: L39Y, L58F, and L39Y/L58F. ITC measurements indicated a significant increase in the affinity to the PRM of variants L58F and L39Y. Measurements taken for the double mutant L39Y/L58F showed the additivity of two mutations leading to about 80-fold increase in the affinity to PRM in comparison to natural protein BLG from bovine milk. The determined crystal structures revealed that pramoxine is accommodated in the β-barrel interior of BLG mutants and stabilized by hydrophobic interactions. The observed additive effect of two mutations on drug binding opens the possibility for further designing of new BLG variants with high affinity to selected drugs.

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β-乳球蛋白变体作为普拉莫辛的潜在载体：全面的结构和生物物理研究。

β-乳球蛋白（BLG）是脂运载蛋白家族的一员。与该组的其他蛋白质一样，BLG可以被修饰以特异性结合医学感兴趣的化合物。本研究的目的是评估L39Y和L58F两个突变在局部麻醉剂吡莫辛（PRM）与β-乳球蛋白结合中的作用。圆二色光谱法、等温滴定量热法（ITC）和X射线晶体学用于了解BLG-PRM相互作用的机制。对三种新的BLG突变体：L39Y、L58F和L39Y/L58F进行了研究。ITC测量表明变体L58F和L39Y对PRM的亲和力显著增加。对双突变体L39Y/L58F进行的测量显示，与来自牛乳的天然蛋白BLG相比，两个突变的相加性导致对PRM的亲和力增加约80倍。所确定的晶体结构表明，吡莫辛被容纳在BLG突变体的β-桶内部，并通过疏水相互作用稳定。观察到的两个突变对药物结合的相加效应为进一步设计对所选药物具有高亲和力的新BLG变体开辟了可能性。

本文章由计算机程序翻译，如有差异，请以英文原文为准。

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来源期刊

Journal of Molecular Recognition 生物-生化与分子生物学

CiteScore

4.60

自引率

3.70%

发文量

审稿时长

2.7 months

期刊介绍： Journal of Molecular Recognition (JMR) publishes original research papers and reviews describing substantial advances in our understanding of molecular recognition phenomena in life sciences, covering all aspects from biochemistry, molecular biology, medicine, and biophysics. The research may employ experimental, theoretical and/or computational approaches. The focus of the journal is on recognition phenomena involving biomolecules and their biological / biochemical partners rather than on the recognition of metal ions or inorganic compounds. Molecular recognition involves non-covalent specific interactions between two or more biological molecules, molecular aggregates, cellular modules or organelles, as exemplified by receptor-ligand, antigen-antibody, nucleic acid-protein, sugar-lectin, to mention just a few of the possible interactions. The journal invites manuscripts that aim to achieve a complete description of molecular recognition mechanisms between well-characterized biomolecules in terms of structure, dynamics and biological activity. Such studies may help the future development of new drugs and vaccines, although the experimental testing of new drugs and vaccines falls outside the scope of the journal. Manuscripts that describe the application of standard approaches and techniques to design or model new molecular entities or to describe interactions between biomolecules, but do not provide new insights into molecular recognition processes will not be considered. Similarly, manuscripts involving biomolecules uncharacterized at the sequence level (e.g. calf thymus DNA) will not be considered.