Comparison of In Silico Signal Sequence-Phospholipid Results with Described In Vitro and In Vivo Protein Translocation Studies Seems to Underscore the Significance of Phospholipids
Abstract
:1. Introduction
2. Results
2.1. Signal Sequence Experimental Results of prePhoE Correlate with In Silico Data
2.2. In Silico Results Suggest a Possible In Silico Protein Translocation Assay
2.3. In Silico Analysis Correlations with In Vitro and In Vivo Findings of Protein Translocation Effects of Signal Sequence Mutations
2.4. In Silico Analysis of Signal Peptide–Lipid Interactions Correlate with In Vitro and In Vivo Findings That Increasing Hydrophobicity in Signal Sequences Changes the Degree of Anionic Phospholipid Dependency in Protein Translocation
2.5. In Vivo Proven Signal Sequence Lipid Contacts in ER Translocation
2.6. An Example of a Closely Related Transmembrane Insertion Experiment
3. Discussion
4. Methods
4.1. Primary and Secondary Structure
4.2. Determination Lipid-Binding Potential and Eisenberg Plot Approach
4.3. Heliquest-Based Eisenberg Plot Approach
4.4. Monte Carlo Simulations Using MCPep
4.5. Structural Modelling
4.6. In Silico Protein Translocation Assay
5. Conclusions
Supplementary Materials
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Name | Sequence | z | <µH> | <H> | LBR a |
---|---|---|---|---|---|
PhoE SP | MKKSTLALVVMGIVASASVQA | 2 | 0.027 | 0.578 | D |
PhoE SP G10L | MKKSTLALVVMLIVASASVQA | 2 | 0.105 | 0.673 | D |
LamB SP | MMITLRKLPLAVAVAAGVMSAQAMA | 2 | 0.222 | 0.582 | D |
LamB SP A13D | MMITLRKLPLAVDVAAGVMSAQAMA | 1 | 0.262 | 0.522 | No |
LamB ∆78 | MMITLRKLP----VAAGVMSAQAMA | 2 | 0.178 | 0.476 | D |
LamB ∆78r1 | MMITLRKLP----VAACVMSAQAMA | 2 | 0.176 | 0.562 | D |
LamB ∆78r2 | MMITLRKLL----VAAGVMSAQAMA | 2 | 0.206 | 0.531 | D |
2K7L SP | MMKKNNLLLLLLLGTANAAS | 2 | 0.165 | 0.637 | D |
2K8L SP | MMKKNNLLLLLLLLGTANAAS | 2 | 0.086 | 0.714 | D |
2K9L SP | MMKKNNLLLLLLLLLGTANAAS | 2 | 0.095 | 0.842 | TM/D |
2K8V SP | MMKKNNVVVVVVVVGTANAAS | 2 | 0.107 | 0.501 | D |
2K9V SP | MMKKNNVVVVVVVVVGTANAAS | 2 | 0.080 | 0.602 | D |
2K10V SP | MMKKNNVVVVVVVVVVGTANAAS | 2 | 0.136 | 0.652 | D |
CLY SP | MRSLLILVLCFLPLAALG | 1 | 0.081 | 1.098 | TM |
CLY R3 | MRRRSLLILVLCFLPLAALG | 3 | 0.100 | 0.918 | TM/D |
CLY R17D | MDSLLILVLCFLPLAALG | −1 | 0.067 | 1.112 | TM |
M13 coat SP | MKKSLVLKASVAVATLVPMLSFA b | 3 | 0.072 | 0.559 | D |
M13 coat SP | MKKSLVLKASVAVATLVPMLSFA b | 1 | 0.109 | 0.863 | TM |
M13 coat SP ∆N | M--------NSVAVATLVPMLSFA | 0 | 0.164 | 0.784 | TM |
M13 coat SP ∆N + 3 | M--------NSLRRRNSVAVATLVPMLSFA | 3 | 0.250 | 0.440 | D |
OmpA SP | MKKTAIAIAVALAGFATVAQA | 2 | 0.251 | 0.620 | D |
OmpA SP I8N | MKKTAIANAVALAGFATVAQA | 2 | 0.118 | 0.487 | D |
OmpA SP L12N | MKKTAIAIAVANAGFATVAQA | 2 | 0.174 | 0.492 | D |
Signal Peptide | Peptide Sequence | Add. AA a |
---|---|---|
PhoE | MKKSTLALVVMGIVASASVQAAEIYNKDGNKLDVYGKV | +17 |
PhoE G10L | MKKSTLALVVMLIVASASVQAAEIYNKDGNKLDVYGKV | +17 |
LamB | MMITLRKLPLAVAVAAGVMSAQAMAVDFHGYARSG | +10 |
LamB A13D | MMITLRKLPLAVDVAAGVMSAQAMA | 0 |
LamB∆78 | MMITLRKLP----VAAGVMSAQAMA | 0 |
LamB∆78r1 | MMITLRKLP----VAACVMSAQAMAVDFH | +4 |
LamB∆78r2 | MMITLRKLL----VAAGVMSAQAMAVDFHGYARSG | +10 |
OmpA | MKKTAIAIAVALAGFATVAQAATSTKKLHKEPATLIKAIDGT | +21 |
OmpA I8N | MKKTAIANAVALAGFATVAQAATSTKKLHKEPATLIKAI | +18 |
OmpA I8D | MKKTAIADAVALAGFATVAQAATSTKKLHKEP | +11 |
OmpA L12N | MKKTAIAIAVANAGFATVAQAATSTKKLHKE | +11 |
OmpA L12D | MKKTAIAIAVADAGFATVAQA | 0 |
2K7L | MMKKNNLLLLLLLGTANAASEIYNKDGNKVDLYGKAV | +17 |
2K8L | MMKKNNLLLLLLLLGTANAASEIYNKDGNKVDLYGKAVGL | +19 |
2K9L | MMKKNNLLLLLLLLLGTANAASEIYNKDGNKVDLYGKAVGL | +19 |
2K8V | MMKKNNVVVVVVVVGTANAASEIYNKDGNKVDLYGK | +15 |
2K9V | MMKKNNVVVVVVVVVGTANAASEIYNKDGNKVDLYGK | +15 |
2K10V | MMKKNNVVVVVVVVVVGTANAASEIYNKDGNKVDLYGKAV | +17 |
CLY | MRSLLILVLCFLPLAALGKVFERCELARTLKRLGMDGYRGIS | +24 |
CLY R3 | MRRRSLLILVLCFLPLAALGKVFERCELARTLKRLGMDGYRGIS | +24 |
CLY R17D | MDSLLILVLCFLPLAALGKVFERCELARTLKRLGMDGYRGIS | +24 |
M13 coat SP | MKKSLVLKASVAVATLVPMLSFAAEGDDPAKAAFNSLQASATEYI | +22 |
M13 coat SP ∆N | M--------NSVAVATLVPMLSFA | 0 |
M13 coat SP ∆N + 3 | M--------NSLRRRNSVAVATLVPMLSFAAEGDDPAKAAFNSL | +14 |
Name | Translocated Mature | ∆Gapp | MCPeppred/ ∆Gapp c | In Vivo/In Vitro Activity |
---|---|---|---|---|
PhoE SP WT | +7 AA (TM) | 1.972 | 100/+ | 100 (+pmf) |
PhoE SP WT | +17 AA (TM) | 1.972 | 100/+ | ND a |
PhoE SP G10L | +7 AA (TM) | 0.958 | 100/++ | 100 (−pmf) |
PhoE SP G10L | +17 AA (TM) | 0.958 | 100/++ | ND |
LamB WT | +10 AA (TM) | 1.633 | 100/+ | 100 |
LamB A13D | +1 AA (Surf.) | 3.818 | 0/− | 10 |
LamB ∆78 | +5 AA (Surf.) b | No pred. | 0/− | 0 |
LamB ∆78r1 | +4 AA (TM) b | 3.798 | 40/− | 50 |
LamB ∆78r2 | +10 AA (TM) b | 2.533 | 100/+ | 90 |
OmpA SP WT | +21 AA (TM) | 1.382 | 100/+ | 100 |
OmpA SP I8N | +18 AA (TM) | 2.581 | 86/− | 0 |
OmpA SP I8D | +11 AA (TM) | 2.933 | 52/− | ND |
OmpA SP L12N | +11 AA (TM) | 2.961 | 52/− | 0 |
OmpA SP L12D | +2 AA (Surf.) | 3.468 | 0/− | ND |
CLY WT | +24 AA | −1.184 | 100/++ | 100 |
CLY R3 | +24 AA | −1.569 | 100/++ | 216 |
CLY R17D | +24 AA | −0.536 | 100/++ | 17 |
M13 SP WT | +22 AA (TM) | 2.806 | 100/+ | 100 |
M13 coat SP ∆N | +3 AA (Surf.) | 3.610 | 0/− | 0 |
M13 coat SP ∆N + 3 | +14 AA (TM) | 4.562 | 64/− | 90 |
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Keller, R.C.A. Comparison of In Silico Signal Sequence-Phospholipid Results with Described In Vitro and In Vivo Protein Translocation Studies Seems to Underscore the Significance of Phospholipids. Lipidology 2024, 1, 3-17. https://doi.org/10.3390/lipidology1010002
Keller RCA. Comparison of In Silico Signal Sequence-Phospholipid Results with Described In Vitro and In Vivo Protein Translocation Studies Seems to Underscore the Significance of Phospholipids. Lipidology. 2024; 1(1):3-17. https://doi.org/10.3390/lipidology1010002
Chicago/Turabian StyleKeller, Rob C. A. 2024. "Comparison of In Silico Signal Sequence-Phospholipid Results with Described In Vitro and In Vivo Protein Translocation Studies Seems to Underscore the Significance of Phospholipids" Lipidology 1, no. 1: 3-17. https://doi.org/10.3390/lipidology1010002