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Accurate and Efficient Resolution of Overlapping Isotopic Envelopes in Protein Tandem Mass Spectra.

Xiao K, Yu F, Fang H, Xue B, Liu Y, Tian Z - Sci Rep (2015)

Bottom Line: The relative deviation (RD) of the overall observed experimental abundance of this OIP relative to the summed ideal value is then calculated.Comprehensive data at the protein and proteome levels, high confidence and good reproducibility were achieved.The resolving method reported here can, in principle, be extended to resolve any envelope-type overlapping data for which the corresponding theoretical reference values are available.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Shanghai Key Laboratory of Chemical Assessment and Sustainability, Tongji University, Shanghai 200092, China.

ABSTRACT
It has long been an analytical challenge to accurately and efficiently resolve extremely dense overlapping isotopic envelopes (OIEs) in protein tandem mass spectra to confidently identify proteins. Here, we report a computationally efficient method, called OIE_CARE, to resolve OIEs by calculating the relative deviation between the ideal and observed experimental abundance. In the OIE_CARE method, the ideal experimental abundance of a particular overlapping isotopic peak (OIP) is first calculated for all the OIEs sharing this OIP. The relative deviation (RD) of the overall observed experimental abundance of this OIP relative to the summed ideal value is then calculated. The final individual abundance of the OIP for each OIE is the individual ideal experimental abundance multiplied by 1 + RD. Initial studies were performed using higher-energy collisional dissociation tandem mass spectra on myoglobin (with direct infusion) and the intact E. coli proteome (with liquid chromatographic separation). Comprehensive data at the protein and proteome levels, high confidence and good reproducibility were achieved. The resolving method reported here can, in principle, be extended to resolve any envelope-type overlapping data for which the corresponding theoretical reference values are available.

No MeSH data available.


Related in: MedlinePlus

The iEF maps of y72-7+ and orthogonal plots of experimental vs. theoretical relative abundance of all interpreted isotopic peaks (with IPAD ≥ 0) without (A,C) and with (B,D) OIE_CARE resolving of OIEs for one of the HCD spectra of myoglobin.The bars and circles in (A,B) are the experimental and theoretical data, respectively. Rel. = relative, Exp. = experimental, and theo. = theoretical.
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f3: The iEF maps of y72-7+ and orthogonal plots of experimental vs. theoretical relative abundance of all interpreted isotopic peaks (with IPAD ≥ 0) without (A,C) and with (B,D) OIE_CARE resolving of OIEs for one of the HCD spectra of myoglobin.The bars and circles in (A,B) are the experimental and theoretical data, respectively. Rel. = relative, Exp. = experimental, and theo. = theoretical.

Mentions: For HCD of myoglobin, IPADs of OIP m/z 1142.617676 in y10-1+ and y72-7+ were reduced from 214 to –2 and 343 to –5, respectively (Table 3). Thus, these two non-MPs were converted into MPs. A total of 141 unique matching b or y ions were found from the three replicate spectra. The number reduced to 134 when the OIE_CARE method was disabled. This implies that 7 more matching b/y ions (b76-7+, y10-1+, y136-13+, y149-15+, y58-6+, y72-7+, y76-7+) were found by resolving OIPs using OIE_CARE. As an example, the iEF maps of y72-7+ without and with using OIE_CARE are shown in Fig. 3(A,B), respectively. The theoretical m/z, theoretical relative abundance, experimental m/z, experimental relative abundance before and after the resolution for each isotopic peak in Fig. 3 are provided in Table S6. It is worth noting that resolving the experimental abundance of OIPs in a tandem mass spectrum increases the number of matching ions (and decreases number of non-matching ones) in general. At the same time, the total number of ions (including both matching and non-matching) remains the same. When a, b, and y ions and their neutral loss (NL) ions were included in the database search, 84.2 ± 0.3% of the interpreted isotopic peaks in myoglobin HCD spectra were found to be OIPs. The overlapping percentage of matching b and y ions was 96.8 ± 0.9%. When a stringent IPMD of 5 ppm was used in the search, the percentages of matching OIPs and overlapping b and y ions were 43.7 ± 2.3% and 60.8 ± 5.3%, respectively. Therefore, the efficient and accurate resolving of such a high percentage of OIPs and OIEs is indispensable to confidently maximizing the matching product ions and protein identification. The OIE_CARE and partition of overlapping abundance of OIPs were used to resolve the experimental relative abundance of all interpreted isotopic peaks with IPAD ≥ 0 from all matching and non-matching ions. The result was that these were also comprehensively brought very close to their corresponding theoretical values. Comparative results from one of the myoglobin HCD spectra with or without using OIE_CARE are presented in Fig. 3(C,D); where the experimental relative abundance of all interpreted isotopic peaks (in all matching and non-matching product ions) are plotted against the corresponding theoretical relative abundance. These abundance together with m/z values are default output of ProteinGoggle for both matching and non-matching product ions. It should be noted that isotopic peaks with IPAD > 0 are, in general, OIPs with a shared experimental abundance. Equivalent plots of the interpreted isotopic peaks with IPAD < 0 show no observed essential changes and are provided in Supplemental Figure S2. As seen from Table 3, to resolve an OIP with n OIEs (or product ions), only 2n + 1 simple arithmetic (addition, subtraction, multiplication, or division) calculations are necessary. This linear computation load relationship with the size of the OIEs is especially advantageous for OIPs with many OIEs. For the HCD spectra of myoglobin, the isotopic peak of m/z is 1123.608521 and is shared by 26 product ions (b111-2H2O-11+, b111-H2O-NH3-11+, b111-2NH3-11+, y142-2H2O-14+, y142-H2O-NH3-14+, b142-2H2O-14+, y142-2NH3-14+, b142-H2O-NH3-14+, a111-11+, b142-2NH3-14+, a92-2H2O-9+, b70-7+, a152-H2O-15+, y10-H2O-1+, y71-7+, b152-2H2O-15+, b152-H2O-NH3-15+, b152-2NH3-15+, y142-H2O-14+, b121-12+, y142-NH3-14+, a71-2H2O-7+, y103-2H2O-10+, a152-15+, and y103-2NH3-10+).


Accurate and Efficient Resolution of Overlapping Isotopic Envelopes in Protein Tandem Mass Spectra.

Xiao K, Yu F, Fang H, Xue B, Liu Y, Tian Z - Sci Rep (2015)

The iEF maps of y72-7+ and orthogonal plots of experimental vs. theoretical relative abundance of all interpreted isotopic peaks (with IPAD ≥ 0) without (A,C) and with (B,D) OIE_CARE resolving of OIEs for one of the HCD spectra of myoglobin.The bars and circles in (A,B) are the experimental and theoretical data, respectively. Rel. = relative, Exp. = experimental, and theo. = theoretical.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4593959&req=5

f3: The iEF maps of y72-7+ and orthogonal plots of experimental vs. theoretical relative abundance of all interpreted isotopic peaks (with IPAD ≥ 0) without (A,C) and with (B,D) OIE_CARE resolving of OIEs for one of the HCD spectra of myoglobin.The bars and circles in (A,B) are the experimental and theoretical data, respectively. Rel. = relative, Exp. = experimental, and theo. = theoretical.
Mentions: For HCD of myoglobin, IPADs of OIP m/z 1142.617676 in y10-1+ and y72-7+ were reduced from 214 to –2 and 343 to –5, respectively (Table 3). Thus, these two non-MPs were converted into MPs. A total of 141 unique matching b or y ions were found from the three replicate spectra. The number reduced to 134 when the OIE_CARE method was disabled. This implies that 7 more matching b/y ions (b76-7+, y10-1+, y136-13+, y149-15+, y58-6+, y72-7+, y76-7+) were found by resolving OIPs using OIE_CARE. As an example, the iEF maps of y72-7+ without and with using OIE_CARE are shown in Fig. 3(A,B), respectively. The theoretical m/z, theoretical relative abundance, experimental m/z, experimental relative abundance before and after the resolution for each isotopic peak in Fig. 3 are provided in Table S6. It is worth noting that resolving the experimental abundance of OIPs in a tandem mass spectrum increases the number of matching ions (and decreases number of non-matching ones) in general. At the same time, the total number of ions (including both matching and non-matching) remains the same. When a, b, and y ions and their neutral loss (NL) ions were included in the database search, 84.2 ± 0.3% of the interpreted isotopic peaks in myoglobin HCD spectra were found to be OIPs. The overlapping percentage of matching b and y ions was 96.8 ± 0.9%. When a stringent IPMD of 5 ppm was used in the search, the percentages of matching OIPs and overlapping b and y ions were 43.7 ± 2.3% and 60.8 ± 5.3%, respectively. Therefore, the efficient and accurate resolving of such a high percentage of OIPs and OIEs is indispensable to confidently maximizing the matching product ions and protein identification. The OIE_CARE and partition of overlapping abundance of OIPs were used to resolve the experimental relative abundance of all interpreted isotopic peaks with IPAD ≥ 0 from all matching and non-matching ions. The result was that these were also comprehensively brought very close to their corresponding theoretical values. Comparative results from one of the myoglobin HCD spectra with or without using OIE_CARE are presented in Fig. 3(C,D); where the experimental relative abundance of all interpreted isotopic peaks (in all matching and non-matching product ions) are plotted against the corresponding theoretical relative abundance. These abundance together with m/z values are default output of ProteinGoggle for both matching and non-matching product ions. It should be noted that isotopic peaks with IPAD > 0 are, in general, OIPs with a shared experimental abundance. Equivalent plots of the interpreted isotopic peaks with IPAD < 0 show no observed essential changes and are provided in Supplemental Figure S2. As seen from Table 3, to resolve an OIP with n OIEs (or product ions), only 2n + 1 simple arithmetic (addition, subtraction, multiplication, or division) calculations are necessary. This linear computation load relationship with the size of the OIEs is especially advantageous for OIPs with many OIEs. For the HCD spectra of myoglobin, the isotopic peak of m/z is 1123.608521 and is shared by 26 product ions (b111-2H2O-11+, b111-H2O-NH3-11+, b111-2NH3-11+, y142-2H2O-14+, y142-H2O-NH3-14+, b142-2H2O-14+, y142-2NH3-14+, b142-H2O-NH3-14+, a111-11+, b142-2NH3-14+, a92-2H2O-9+, b70-7+, a152-H2O-15+, y10-H2O-1+, y71-7+, b152-2H2O-15+, b152-H2O-NH3-15+, b152-2NH3-15+, y142-H2O-14+, b121-12+, y142-NH3-14+, a71-2H2O-7+, y103-2H2O-10+, a152-15+, and y103-2NH3-10+).

Bottom Line: The relative deviation (RD) of the overall observed experimental abundance of this OIP relative to the summed ideal value is then calculated.Comprehensive data at the protein and proteome levels, high confidence and good reproducibility were achieved.The resolving method reported here can, in principle, be extended to resolve any envelope-type overlapping data for which the corresponding theoretical reference values are available.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Shanghai Key Laboratory of Chemical Assessment and Sustainability, Tongji University, Shanghai 200092, China.

ABSTRACT
It has long been an analytical challenge to accurately and efficiently resolve extremely dense overlapping isotopic envelopes (OIEs) in protein tandem mass spectra to confidently identify proteins. Here, we report a computationally efficient method, called OIE_CARE, to resolve OIEs by calculating the relative deviation between the ideal and observed experimental abundance. In the OIE_CARE method, the ideal experimental abundance of a particular overlapping isotopic peak (OIP) is first calculated for all the OIEs sharing this OIP. The relative deviation (RD) of the overall observed experimental abundance of this OIP relative to the summed ideal value is then calculated. The final individual abundance of the OIP for each OIE is the individual ideal experimental abundance multiplied by 1 + RD. Initial studies were performed using higher-energy collisional dissociation tandem mass spectra on myoglobin (with direct infusion) and the intact E. coli proteome (with liquid chromatographic separation). Comprehensive data at the protein and proteome levels, high confidence and good reproducibility were achieved. The resolving method reported here can, in principle, be extended to resolve any envelope-type overlapping data for which the corresponding theoretical reference values are available.

No MeSH data available.


Related in: MedlinePlus