TY - JOUR
T1 - Biochemical and spectroscopic characterization of dinuclear Mn-sites in artificial four-helix bundle proteins
AU - Olson, Tien L.
AU - Espiritu, Eduardo
AU - Edwardraja, Selvakumar
AU - Canarie, Elizabeth
AU - Flores, Marco
AU - Williams, Joann
AU - Ghirlanda, Giovanna
AU - Allen, James
N1 - Funding Information:
This work was supported by funding from the National Science Foundation ( CHE 1505874 ).
Publisher Copyright:
© 2017 Elsevier B.V.
PY - 2017/12
Y1 - 2017/12
N2 - To better understand metalloproteins with Mn-clusters, we have designed artificial four-helix bundles to have one, two, or three dinuclear metal centers able to bind Mn(II). Circular dichroism measurements showed that the Mn-proteins have substantial α-helix content, and analysis of electron paramagnetic resonance spectra is consistent with the designed number of bound Mn-clusters. The Mn-proteins were shown to catalyze the conversion of hydrogen peroxide into molecular oxygen. The loss of hydrogen peroxide was dependent upon the concentration of protein with bound Mn, with the proteins containing multiple Mn-clusters showing greater activity. Using an oxygen sensor, the oxygen concentration was found to increase with a rate up to 0.4 μM/min, which was dependent upon the concentrations of hydrogen peroxide and the Mn-protein. In addition, the Mn-proteins were shown to serve as electron donors to bacterial reaction centers using optical spectroscopy. Similar binding of the Mn-proteins to reaction centers was observed with an average dissociation constant of 2.3 μM. The Mn-proteins with three metal centers were more effective at this electron transfer reaction than the Mn-proteins with one or two metal centers. Thus, multiple Mn-clusters can be incorporated into four-helix bundles with the capability of performing catalysis and electron transfer to a natural protein.
AB - To better understand metalloproteins with Mn-clusters, we have designed artificial four-helix bundles to have one, two, or three dinuclear metal centers able to bind Mn(II). Circular dichroism measurements showed that the Mn-proteins have substantial α-helix content, and analysis of electron paramagnetic resonance spectra is consistent with the designed number of bound Mn-clusters. The Mn-proteins were shown to catalyze the conversion of hydrogen peroxide into molecular oxygen. The loss of hydrogen peroxide was dependent upon the concentration of protein with bound Mn, with the proteins containing multiple Mn-clusters showing greater activity. Using an oxygen sensor, the oxygen concentration was found to increase with a rate up to 0.4 μM/min, which was dependent upon the concentrations of hydrogen peroxide and the Mn-protein. In addition, the Mn-proteins were shown to serve as electron donors to bacterial reaction centers using optical spectroscopy. Similar binding of the Mn-proteins to reaction centers was observed with an average dissociation constant of 2.3 μM. The Mn-proteins with three metal centers were more effective at this electron transfer reaction than the Mn-proteins with one or two metal centers. Thus, multiple Mn-clusters can be incorporated into four-helix bundles with the capability of performing catalysis and electron transfer to a natural protein.
KW - Catalase
KW - De novo design
KW - Electron transfer
KW - Metalloproteins
KW - Photosynthesis
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U2 - 10.1016/j.bbabio.2017.08.013
DO - 10.1016/j.bbabio.2017.08.013
M3 - Article
C2 - 28882760
AN - SCOPUS:85029611397
SN - 0005-2728
VL - 1858
SP - 945
EP - 954
JO - Biochimica et Biophysica Acta - Bioenergetics
JF - Biochimica et Biophysica Acta - Bioenergetics
IS - 12
ER -