biostudies-literatureYang JNatural Science Foundation of Beijing MunicipalityMinistry of Science and Technology of the People&apos;s Republic of ChinaNational Natural Science Foundation of China8302-8324https://www.ebi.ac.uk/biostudies/studies/S-EPMC7294093biostudies-literatureUnknown295(24)Heat shock protein 70 (Hsp70) proteins are a family of ancient and conserved chaperones. Cysteine modifications have been widely detected among different Hsp70 family members <i>in vivo</i>, but their effects on Hsp70 structure and function are unclear. Here, we treated HeLa cells with diamide, which typically induces disulfide bond formation except in the presence of excess GSH, when glutathionylated cysteines predominate. We show that in these cells, HspA1A (hHsp70) undergoes reversible cysteine modifications, including glutathionylation, potentially at all five cysteine residues. <i>In vitro</i> experiments revealed that modification of cysteines in the nucleotide-binding domain of hHsp70 is prevented by nucleotide binding but that Cys-574 and Cys-603, located in the C-terminal α-helicaThe Journal of biological chemistry<i>S</i>-Glutathionylation of human inducible Hsp70 reveals a regulatory mechanism involving the C-terminal α-helical lid.PMC729409321673278315708573157078031770829517202691849203314707472017YFA050400031920103011Yang JLiu ZHu WZhang HWu SChen XChen CPerrett SGong WWu HWang Lfalse<i>S</i>-Glutathionylation of human inducible Hsp70 reveals a regulatory mechanism involving the C-terminal α-helical lid.Heat shock protein 70 (Hsp70) proteins are a family of ancient and conserved chaperones. Cysteine modifications have been widely detected among different Hsp70 family members <i>in vivo</i>, but their effects on Hsp70 structure and function are unclear. Here, we treated HeLa cells with diamide, which typically induces disulfide bond formation except in the presence of excess GSH, when glutathionylated cysteines predominate. We show that in these cells, HspA1A (hHsp70) undergoes reversible cysteine modifications, including glutathionylation, potentially at all five cysteine residues. <i>In vitro</i> experiments revealed that modification of cysteines in the nucleotide-binding domain of hHsp70 is prevented by nucleotide binding but that Cys-574 and Cys-603, located in the C-terminal α-helica2020-01-01T00:00:00Z2020 Jun2026-04-30T05:57:00.235Z2020-06-25T07:29:10ZS-EPMC72940933233210110.1074/jbc.RA119.012372