Complete Charge Regulation by a Redox Enzyme Upon Single Electron Transfer

Zhang, Ao Yun; Koone, Jordan C.; Dashnaw, Chad M.; Zahler, Collin T.; Shaw, Bryan F.

doi:10.1002/ange.202001452

Cited by 1 publication

(4 citation statements)

References 43 publications

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“…61 In general, charge regulation might be an evolutionarily adopted property by which proteins maintain Z during ligand binding 62 or catalysis. 63 For example, we have already shown that superoxide dismutase maintains tight regulation of its Z during single electron transfer (and fluctuations in pH). We hypothesize that this property is significant to SOD1's ability to bind superoxide independent of its copper oxidation state at various subcellular pH values.…”

Section: Resultsmentioning

confidence: 99%

“…We hypothesize that this property is significant to SOD1's ability to bind superoxide independent of its copper oxidation state at various subcellular pH values. 63,64 Examining charge regulation during crowding of proteins that do or do not regulate their charge upon ΔpH or ET is an area of interest.…”

Section: Resultsmentioning

confidence: 99%

“…In general, charge regulation might be an evolutionarily adopted property by which proteins maintain Z during ligand binding 62 or catalysis 63 . For example, we have already shown that superoxide dismutase maintains tight regulation of its Z during single electron transfer (and fluctuations in pH).…”

Section: Resultsmentioning

confidence: 99%

“…For example, we have already shown that superoxide dismutase maintains tight regulation of its Z during single electron transfer (and fluctuations in pH). We hypothesize that this property is significant to SOD1's ability to bind superoxide independent of its copper oxidation state at various subcellular pH values 63,64 . Examining charge regulation during crowding of proteins that do or do not regulate their charge upon ΔpH or ET is an area of interest.…”

Section: Resultsmentioning

confidence: 99%

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Measuring how two proteins affect each other's net charge in a crowded environment

et al. 2021

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Theory predicts that the net charge (Z) of a protein can be altered by the net charge of a neighboring protein as the two approach one another below the Debye length. This type of charge regulation suggests that a protein's charge and perhaps function might be affected by neighboring proteins without direct binding. Charge regulation during protein crowding has never been directly measured due to analytical challenges. Here, we show that lysine specific protein crosslinkers (NHS ester-Staudinger pairs) can be used to mimic crowding by linking two non-interacting proteins at a maximal distance of $7.9 Å. The net charge of the regioisomeric dimers and preceding monomers can then be determined with lysine-acyl "protein charge ladders" and capillary electrophoresis. As a proof of concept, we covalently linked myoglobin (Z monomer = À0.43 ± 0.01) and α-lactalbumin (Z monomer = À4.63 ± 0.05). Amide hydrogen/deuterium exchange and circular dichroism spectroscopy demonstrated that crosslinking did not significantly alter the structure of either protein or result in direct binding (thus mimicking crowding). Ultimately, capillary electrophoretic analysis of the dimeric charge ladder detected a change in charge of ΔZ = À0.04 ± 0.09 upon crowding by this pair (Z dimer = À5.10 ± 0.07). These small values of ΔZ are not necessarily general to protein crowding (qualitatively or quantitatively) but will vary per protein size, charge, and solvent conditions.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%