Start2Fold

The database of hydrogen/deuterium exchange data on protein folding and stability

Entry STF0010

Dictyostelium discoideum hisactophilin-1

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Protein information

Name of the protein: Hisactophilin-1
Organism: Dictyostelium discoideum (Slime mold)
Number of residues: 118
Related UniProt entry:   P13231 (Fragment: 1 - 118)
Related PDB entry:   1HCE

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Experiment sets

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EARLY

Method: Quenched-flow HDX NMR

Conditions: pH 7.8; 20.0 Celsius; Probes: 31

Related publication:
 PMID 11847289

Experiment details: "Denatured perdeutero hisactophilin (8.0 mg/mL in 6.4 M urea-D4, 250 mM potassium phosphate buffer at pH 7.8, 5 mM DTT, 5 mM EDTA, D2O) was diluted fivefold with D2O, incubated for a series of set refolding times, pH-pulsed using 5 volumes of 62.5 mM glycine/sodium glycinate in H2O (final pH was 9.52 for 29 ms), and pH-quenched with 4 volumes of 112.5 mM triethanolamine buffer in H2O (final pH was 7.7). The sample was immediately concentrated and exchanged by ultrafiltration into 50 mM triethanolamine buffer at pH 7.8 in D2O. Final sample volumes (0.5 mL) contained 5mg/mL hisactophilin. NMR experiments were performed within 2 h after concentration."

Protection threshold: Fast protection rate s(-1) > 20

Sequence: MGNRAFKSHHGHFLSAEGEAVKTHHGHHDHHTHFHVENHGGKVALKTHCGKYLSIGDHKQVYLSHHLHGDHSLFHLEHHGGKVSIKGHHHHYISADHHGHVSTKEHHDHDTTFEEIII
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EARLY residues

14: L; 43: V; 44: A; 45: L; 47: T; 53: L; 62: Y; 73: L; 74: F; 114: E;
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INTERMEDIATE

Method: Quenched-flow HDX NMR

Conditions: pH 7.8; 20.0 Celsius; Probes: 31

Related publication:
 PMID 11847289

Experiment details: "Denatured perdeutero hisactophilin (8.0 mg/mL in 6.4 M urea-D4, 250 mM potassium phosphate buffer at pH 7.8, 5 mM DTT, 5 mM EDTA, D2O) was diluted fivefold with D2O, incubated for a series of set refolding times, pH-pulsed using 5 volumes of 62.5 mM glycine/sodium glycinate in H2O (final pH was 9.52 for 29 ms), and pH-quenched with 4 volumes of 112.5 mM triethanolamine buffer in H2O (final pH was 7.7). The sample was immediately concentrated and exchanged by ultrafiltration into 50 mM triethanolamine buffer at pH 7.8 in D2O. Final sample volumes (0.5 mL) contained 5mg/mL hisactophilin. NMR experiments were performed within 2 h after concentration."

Protection threshold: 10 < fast protection rate s(-1) < 20

Sequence: MGNRAFKSHHGHFLSAEGEAVKTHHGHHDHHTHFHVENHGGKVALKTHCGKYLSIGDHKQVYLSHHLHGDHSLFHLEHHGGKVSIKGHHHHYISADHHGHVSTKEHHDHDTTFEEIII
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INTERMEDIATE residues

15: S; 34: F; 35: H; 46: K; 54: S; 55: I; 83: V; 84: S; 93: I;
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LATE

Method: Quenched-flow HDX NMR

Conditions: pH 7.8; 20.0 Celsius; Probes: 31

Related publication:
 PMID 11847289

Experiment details: "Denatured perdeutero hisactophilin (8.0 mg/mL in 6.4 M urea-D4, 250 mM potassium phosphate buffer at pH 7.8, 5 mM DTT, 5 mM EDTA, D2O) was diluted fivefold with D2O, incubated for a series of set refolding times, pH-pulsed using 5 volumes of 62.5 mM glycine/sodium glycinate in H2O (final pH was 9.52 for 29 ms), and pH-quenched with 4 volumes of 112.5 mM triethanolamine buffer in H2O (final pH was 7.7). The sample was immediately concentrated and exchanged by ultrafiltration into 50 mM triethanolamine buffer at pH 7.8 in D2O. Final sample volumes (0.5 mL) contained 5mg/mL hisactophilin. NMR experiments were performed within 2 h after concentration."

Protection threshold: 4 < fast protection rate s(-1) < 10

Sequence: MGNRAFKSHHGHFLSAEGEAVKTHHGHHDHHTHFHVENHGGKVALKTHCGKYLSIGDHKQVYLSHHLHGDHSLFHLEHHGGKVSIKGHHHHYISADHHGHVSTKEHHDHDTTFEEIII
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LATE residues

5: A; 6: F; 7: K; 8: S; 63: L; 86: K; 87: G; 92: Y; 94: S; 95: A; 113: F;
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STRONG

Method: Native exchange NMR

Conditions: pH 5.9-9.7; 17.4 Celsius; Probes: 37

Related publication:
 PMID 11802717

Experiment details: "For measurement of the pH dependence of amide exchange rates, purified protein was exchanged into 50 mM potassium phosphate buffer at pH 5.9, 6.8, and 7.8 or 50 mM glycine buffer at pH 8.7 and 9.7 and then lyophilized. H/D exchange was initiated by redissolving the sample in D2O to the same volume that existed prior to lyophilization. The slow exchange core is defined by the authors as the collection of residues with the slowest average exchange rate, the strongest dependence of exchange on denaturant concentration, and the strongest temperature dependence for exchange rates."

Protection threshold: Slow exchange core

Sequence: MGNRAFKSHHGHFLSAEGEAVKTHHGHHDHHTHFHVENHGGKVALKTHCGKYLSIGDHKQVYLSHHLHGDHSLFHLEHHGGKVSIKGHHHHYISADHHGHVSTKEHHDHDTTFEEIII
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STRONG residues

44: A; 45: L; 46: K; 54: S; 62: Y; 74: F; 85: I; 86: K; 93: I; 94: S; 113: F; 114: E;
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