Dual role of the active-center cysteine in human peroxiredoxin 1: Peroxidase activity and heme binding

Abstract

HBP23, a 23-kDa heme-binding protein identified in rats, is a member of the peroxiredoxin (Prx) family, the primary peroxidases involved in hydrogen peroxide catabolism. Although HBP23 has a characteristic Cys-Pro heme-binding motif, the significance of heme binding to Prx family proteins remains to be elucidated. Here, we examined the effect of heme binding to human peroxiredoxin-1 (PRX1), which has 97% amino acid identity to HBP23. PRX1 was expressed in Escherichia coli and purified to homogeneity. Spectroscopic titration demonstrated that PRX1 binds heme with a 1:1 stoichiometry and a dissociation constant of 0.17 μM. UV-vis spectra of heme-PRX1 suggested that Cys52 is the axial ligand of ferric heme. PRX1 peroxidase activity was lost upon heme binding, reflecting the fact that Cys52 is not only the heme-binding site but also the active center of peroxidase activity. Interestingly, heme binding to PRX1 caused a decrease in the toxicity and degradation of heme, significantly suppressing H 2 O 2 -dependent heme peroxidase activity and degradation of PRX1-bound heme compared with that of free hemin. By virtue of its cytosolic abundance (~20 μM), PRX1 thus functions as a scavenger of cytosolic hemin (<1 μM). Collectively, our results indicate that PRX1 has a dual role; Cys-dependent peroxidase activity and cytosolic heme scavenger.

The Iron Chaperone Protein CyaY from Vibrio cholerae Is a Heme-Binding Protein

“…This peak position and the shape of the Soret band were slightly, but detectably, different from those of free hemin (λ max , ~387 nm), indicating that heme specifically binds to VcCyaY ( Biochemistry 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 13 Q-bands at 507 and 619 nm ( Figure 2C). This spectrum was similar to those of heme-bound Bach1 (371 nm) 29 and peroxiredoxin 1 (370 nm) 30 , in which cysteine serves as a heme ligand.…”

Handling heme: The mechanisms underlying the movement of heme within and between cells

“…An illustrative but interesting case is that of HBP23/PRX1. PRX1 is a thiol peroxidase that was recently found to bind heme with a 1:1 stoichiometry and a dissociation constant of 170 nM [69], which is within the 20-340 nM consensus range of labile heme in cells [21][22][23]. PRX1, which binds heme using a catalytic Cys, Cys52, exhibits a loss in thiol peroxidase activity upon heme binding.…”

“…PRX1, which binds heme using a catalytic Cys, Cys52, exhibits a loss in thiol peroxidase activity upon heme binding. [69] Additionally, heme binding to PRX1 suppresses both the peroxidase activity of heme and the H 2 O 2-mediated degradation of heme. [69] Given the abundance of PRX1 in the cytosol, 20 μM, and ability to bind and potentially detoxify and protect heme from degradation, PRX1 may have dual roles as a peroxide scavenging thiol peroxidase and a heme buffering or storage protein.…”

“…[69] Additionally, heme binding to PRX1 suppresses both the peroxidase activity of heme and the H 2 O 2-mediated degradation of heme. [69] Given the abundance of PRX1 in the cytosol, 20 μM, and ability to bind and potentially detoxify and protect heme from degradation, PRX1 may have dual roles as a peroxide scavenging thiol peroxidase and a heme buffering or storage protein. However, as with many other heme binding proteins proposed to play a role in heme homeostasis, the in vivo function of PRX1 in heme homeostasis has yet to be demonstrated [69].…”

Biological and Chemical Effects by Endo- And Exogenous Hydrogen Peroxide in Human Body Cells Structures

“…Известно, что для выполнения фагоци-тарной функции эффекторными иммуноци-тами необходим каскад реакций с выделе-нием hydrogen peroxide [12,24]. Макрофаги способствуют поддержанию гомеостаза, очищая организм от стареющих и апопто-тических клеток, восстанавливая ткани по-сле инфекции и травмы, выполняя ведущую роль в защите организма.…”