Effect of terminal phosphate groups on collisional dissociation of RNA oligonucleotide anions
ABSTRACT An increasing need of mass spectrometric analysis of RNA molecules calls for a better understanding of their gas-phase fragmentation behaviors. In this study, we investigated the effect of terminal phosphate groups on the fragmentation spectra of RNA oligonucleotides (oligos) using high-resolution mass spectrometry (MS). Negative-ion mode collision-induced dissociation (CID) and higher-energy collisional dissociation (HCD) were carried out on RNA oligos containing a terminal phosphate group on either or both ends, or neither. We find that terminal phosphate groups affect the fragmentation behavior of RNA oligos in a way that depends on the precursor charge state and the oligo length. Specifically, for precursor ions of RNA oligos of the same sequence, those with 5’- or 3’-phosphate, or both, have a higher charge state distribution and lose upon CID or HCD the phosphate group(s) in the form of a neutral (H3PO4or HPO3) or an anion ([H2PO4]-or [PO3]-). Such neutral or charged loss is most conspicuous for precursor ions of an intermediate charge state, e.g. 3-for 4-nt oligos or 4-and 5-for 8-nt oligos. This decreases the intensity of sequencing ions (a-, a-B, b-, c-, d-, w-, x-, y-, z-ions), hence unfavorable for sequencing by CID or HCD. Removal of terminal phosphate groups by calf intestinal alkaline phosphatase improved MS analysis of RNA oligos. Additionally, the intensity of a fragment ion atm/z158.925, which we have identified as a dehydrated pyrophosphate anion ([HP2O6]-), is markedly increased by the presence of a terminal phosphate group. These findings expand the knowledge base necessary for software development for MS analysis of RNA..
| Medienart: |
|
|---|
Preprint| Erscheinungsjahr: |
2024 |
|---|---|
| Erschienen: |
2024 |
| Enthalten in: |
bioRxiv.org - (2024) vom: 21. Apr. Zur Gesamtaufnahme - year:2024 |
|---|
| Sprache: |
Englisch |
|---|
| Beteiligte Personen: |
Zuo, Mei-Qing [VerfasserIn] |
|---|
| Links: |
Volltext [kostenfrei] |
|---|
| Themen: |
|---|
| doi: |
10.1101/2024.03.05.583607 |
|---|
| funding: |
|
|---|---|
| Förderinstitution / Projekttitel: |
|
| PPN (Katalog-ID): |
XBI042831946 |
|---|
| LEADER | 01000caa a22002652 4500 | ||
|---|---|---|---|
| 001 | XBI042831946 | ||
| 003 | DE-627 | ||
| 005 | 20240421090536.0 | ||
| 007 | cr uuu---uuuuu | ||
| 008 | 240308s2024 xx |||||o 00| ||eng c | ||
| 024 | 7 | |a 10.1101/2024.03.05.583607 |2 doi | |
| 035 | |a (DE-627)XBI042831946 | ||
| 035 | |a (biorXiv)10.1101/2024.03.05.583607 | ||
| 040 | |a DE-627 |b ger |c DE-627 |e rakwb | ||
| 041 | |a eng | ||
| 100 | 1 | |a Zuo, Mei-Qing |e verfasserin |4 aut | |
| 245 | 1 | 0 | |a Effect of terminal phosphate groups on collisional dissociation of RNA oligonucleotide anions |
| 264 | 1 | |c 2024 | |
| 336 | |a Text |b txt |2 rdacontent | ||
| 337 | |a Computermedien |b c |2 rdamedia | ||
| 338 | |a Online-Ressource |b cr |2 rdacarrier | ||
| 520 | |a ABSTRACT An increasing need of mass spectrometric analysis of RNA molecules calls for a better understanding of their gas-phase fragmentation behaviors. In this study, we investigated the effect of terminal phosphate groups on the fragmentation spectra of RNA oligonucleotides (oligos) using high-resolution mass spectrometry (MS). Negative-ion mode collision-induced dissociation (CID) and higher-energy collisional dissociation (HCD) were carried out on RNA oligos containing a terminal phosphate group on either or both ends, or neither. We find that terminal phosphate groups affect the fragmentation behavior of RNA oligos in a way that depends on the precursor charge state and the oligo length. Specifically, for precursor ions of RNA oligos of the same sequence, those with 5’- or 3’-phosphate, or both, have a higher charge state distribution and lose upon CID or HCD the phosphate group(s) in the form of a neutral (H3PO4or HPO3) or an anion ([H2PO4]-or [PO3]-). Such neutral or charged loss is most conspicuous for precursor ions of an intermediate charge state, e.g. 3-for 4-nt oligos or 4-and 5-for 8-nt oligos. This decreases the intensity of sequencing ions (a-, a-B, b-, c-, d-, w-, x-, y-, z-ions), hence unfavorable for sequencing by CID or HCD. Removal of terminal phosphate groups by calf intestinal alkaline phosphatase improved MS analysis of RNA oligos. Additionally, the intensity of a fragment ion atm/z158.925, which we have identified as a dehydrated pyrophosphate anion ([HP2O6]-), is markedly increased by the presence of a terminal phosphate group. These findings expand the knowledge base necessary for software development for MS analysis of RNA. | ||
| 650 | 4 | |a Biology |7 (dpeaa)DE-84 | |
| 650 | 4 | |a 570 |7 (dpeaa)DE-84 | |
| 700 | 1 | |a Song, Ge |e verfasserin |4 aut | |
| 700 | 1 | |a Dong, Meng-Qiu |e verfasserin |0 (orcid)0000-0002-6094-1182 |4 aut | |
| 700 | 1 | |a Sun, Rui-Xiang |e verfasserin |4 aut | |
| 773 | 0 | 8 | |i Enthalten in |t bioRxiv.org |g (2024) vom: 21. Apr. |
| 773 | 1 | 8 | |g year:2024 |g day:21 |g month:04 |
| 856 | 4 | 0 | |u http://dx.doi.org/10.1101/2024.03.05.583607 |x 0 |z kostenfrei |3 Volltext |
| 912 | |a GBV_XBI | ||
| 951 | |a AR | ||
| 952 | |j 2024 |b 21 |c 04 | ||