Biophysical and NMR analysis reveals binding affinity between HAX1 and CLPB proteins  

作　　者：Huiqin Zhang Yong Liu Yunyan Li Maosen Ruan Shu Zhou Junfeng Wang Jing Yang 

机构地区：[1]Institute of Physical Science and Information Technology,Anhui University,Hefei,230601,China [2]High Magnetic Field Laboratory,Key Laboratory of High Magnetic Field and Ion Beam Physical Biology,Hefei Institutes of Physical Science,Chinese Academy of Sciences,Hefei,230031,China [3]University of Science and Technology of China,Hefei,230026,China [4]International Magnetobiology Frontier Research Center,Hefei,230031,China

出　　处：《Magnetic Resonance Letters》2025年第1期12-21,共10页磁共振快报(英文)

基　　金：supported by grants from the Special Foundation of President of the Chinese Academy of Sciences(Grant No.,YZJJ2020QN27,YZJJ2021QN33);Anhui Provincial Natural Science Foundation(Grant No.,2108085MC79).

摘　　要：HCLS1-associated protein X-1(HAX1)is a multifunctional mitochondrial protein involved in the regulation of apoptosis,a crucial process of programmed cell death,and mRNA processing.Despite its significance,limited structural data is available for HAX1,hindering a comprehensive understanding of its biological function.Notably,the caseinolytic mitochondrial matrix peptidase chaperone subunit B(CLPB)has been identified as an interacting partner of HAX1,yet the biophysical properties and binding affinity governing their interaction remain poorly defined.In this study,we present a thorough biophysical characterization of full-length human HAX1 and CLPB,accomplished through recombinant expression and purification.By employing size exclusion chromatography,dynamic light scattering,and circular dichroism spectroscopy,we successfully established their biophysical properties,revealing contrasting structural features,with CLPB displaying a-helical content and HAX1 exhibiting a disordered nature.Moreover,we employed solutionstate nuclear magnetic resonance(NMR)spectroscopy to probe their binding affinity.Our findings demonstrate the formation of stable multimeric complexes between HAX1 and CLPB,and we quantified a dissociation constant in the low range of micro-molar for their high affinity interaction.These results lay the foundation for further in-depth investigations into the dynamics and energetics governing the HAX1-CLPB interaction,ultimately contributing to a comprehensive understanding of their functional mechanisms.

关 键 词：HAX1 CLPB Protein interaction Biophysical characterization NMR spectroscopy 

分 类 号：R73[医药卫生—肿瘤]