Abstract
Native electron capture dissociation (NECD) is a process during which proteins undergo fragmentation similar to that from radical dissociation methods, but without the addition of exogenous electrons. However, after three initial reports of NECD from the cytochrome c dimer complex, no further evidence of the effect has been published. Here, we report NECD behavior from horse spleen ferritin, a ∼490 kDa protein complex ∼20-fold larger than the previously studied cytochrome c dimer. Application of front-end infrared excitation (FIRE) in conjunction with low- and high-m/z quadrupole isolation and collisionally activated dissociation (CAD) provides new insights into the NECD mechanism. Additionally, activation of the intact complex in either the electrospray droplet or the gas phase produced c-type fragment ions. Similar to the previously reported results on cytochrome c, these fragment ions form near residues known to interact with iron atoms in solution. By mapping the location of backbone cleavages associated with c-type ions onto the crystal structure, we are able to characterize two distinct iron binding channels that facilitate iron ion transport into the core of the complex. The resulting pathways are in good agreement with previously reported results for iron binding sites in mammalian ferritin. (Graph Presented).
| Original language | English |
|---|---|
| Pages (from-to) | 10711-10716 |
| Number of pages | 6 |
| Journal | Analytical Chemistry |
| Volume | 89 |
| Issue number | 20 |
| DOIs | |
| Publication status | Published - Oct 17 2017 |
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ASJC Scopus subject areas
- Analytical Chemistry
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Native Electron Capture Dissociation Maps to Iron-Binding Channels in Horse Spleen Ferritin. / Skinner, Owen S.; McAnally, Michael O.; Van Duyne, Richard P.; Schatz, George C; Breuker, Kathrin; Compton, Philip D.; Kelleher, Neil L.
In: Analytical Chemistry, Vol. 89, No. 20, 17.10.2017, p. 10711-10716.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Native Electron Capture Dissociation Maps to Iron-Binding Channels in Horse Spleen Ferritin
AU - Skinner, Owen S.
AU - McAnally, Michael O.
AU - Van Duyne, Richard P.
AU - Schatz, George C
AU - Breuker, Kathrin
AU - Compton, Philip D.
AU - Kelleher, Neil L.
PY - 2017/10/17
Y1 - 2017/10/17
N2 - Native electron capture dissociation (NECD) is a process during which proteins undergo fragmentation similar to that from radical dissociation methods, but without the addition of exogenous electrons. However, after three initial reports of NECD from the cytochrome c dimer complex, no further evidence of the effect has been published. Here, we report NECD behavior from horse spleen ferritin, a ∼490 kDa protein complex ∼20-fold larger than the previously studied cytochrome c dimer. Application of front-end infrared excitation (FIRE) in conjunction with low- and high-m/z quadrupole isolation and collisionally activated dissociation (CAD) provides new insights into the NECD mechanism. Additionally, activation of the intact complex in either the electrospray droplet or the gas phase produced c-type fragment ions. Similar to the previously reported results on cytochrome c, these fragment ions form near residues known to interact with iron atoms in solution. By mapping the location of backbone cleavages associated with c-type ions onto the crystal structure, we are able to characterize two distinct iron binding channels that facilitate iron ion transport into the core of the complex. The resulting pathways are in good agreement with previously reported results for iron binding sites in mammalian ferritin. (Graph Presented).
AB - Native electron capture dissociation (NECD) is a process during which proteins undergo fragmentation similar to that from radical dissociation methods, but without the addition of exogenous electrons. However, after three initial reports of NECD from the cytochrome c dimer complex, no further evidence of the effect has been published. Here, we report NECD behavior from horse spleen ferritin, a ∼490 kDa protein complex ∼20-fold larger than the previously studied cytochrome c dimer. Application of front-end infrared excitation (FIRE) in conjunction with low- and high-m/z quadrupole isolation and collisionally activated dissociation (CAD) provides new insights into the NECD mechanism. Additionally, activation of the intact complex in either the electrospray droplet or the gas phase produced c-type fragment ions. Similar to the previously reported results on cytochrome c, these fragment ions form near residues known to interact with iron atoms in solution. By mapping the location of backbone cleavages associated with c-type ions onto the crystal structure, we are able to characterize two distinct iron binding channels that facilitate iron ion transport into the core of the complex. The resulting pathways are in good agreement with previously reported results for iron binding sites in mammalian ferritin. (Graph Presented).
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U2 - 10.1021/acs.analchem.7b01581
DO - 10.1021/acs.analchem.7b01581
M3 - Article
C2 - 28938074
AN - SCOPUS:85031507684
VL - 89
SP - 10711
EP - 10716
JO - Analytical Chemistry
JF - Analytical Chemistry
SN - 0003-2700
IS - 20
ER -