1. The A20I mutant has the strongest ice-binding ability, and the order of ice-growth inhibition strength is A20L < A20G < WT < A20T < A20V < A20I.
2. The location of the γ-CH3 group at the 20th residue plays a crucial role in the ice-binding ability of this IBP to the ice planes.
AFP013000
General Information
| Protein Name | Ice-structuring protein |
| UniProt ID | Q53UJ4 |
| Nucleotide Sequence ID in NCBI | AB188394 |
| Protein Sequence ID in NCBI | BAD95781.1 |
| Species | Zoarces elongatus (Zoarces elongatus Kner) (Notched-fin eelpout, NFE) |
| Sequence Length | 67 |
| Sequence | |
| Structure | PDB ID: 5xqn |
| Solvent Accessible Surface Area | Total SASA | 4077.44 Ų | Polar SASA | 2516.96 Ų | Apolar SASA | 6594.40 Ų |
AFP013000000
| Mutation | Wild Type |
| Sequence |
Ice crystal morphology
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (wild type) |
| Ice crystal morphology | Fig.2.A Comparison of the strength of ice-growth inhibition between WT andmutant proteins. (A) Images of bipyramidal ice crystals captured for A20L, A20G,WT, A20T, A20V, and A20I mutants before and after 9 min of annealing time. Thearrow indicates the lengthof the a axis. |
Ice crystal morphology
| PMID | 30760774 |
| DOI | 10.1038/s41598-018-36546-2 |
| Protein Name | Zoarces elongatus AFPIII |
| Ice crystal morphology | Fig.4.B Morphology of ice crystals observed for AFPI-III, A20L, and Tis8 at concentrations lower than their IRI endpoint. The hexagonal ice plate is created in the solutions of AFPI-AFPIII and A20L, while it is not facetted for Tis8 and similarly for the solvent (40% sucrose). |
Ice crystal morphology
| PMID | 19187223 |
| DOI | 10.1111/j.1742-4658.2009.06887.x |
| Protein Name | nfeAFP6 (SP) |
| Ice crystal morphology | Fig.1 Morphological change in an ice crystal observed for 0.1 mM solutions of nfeAFP8 (QAE isoform) and nfeAFP6 (SP isoform). (A) Photomicroscope images for nfeAFP8 (a-d) and nfeAFP6 (e-h) were obtained at a cooling rate of 0.20 ℃·min-1. (a, e) T = Tm - 0.1 ℃; (b, f) T = Tm -0.2 ℃; (c, g) T = Tburst; (d, h) T = Tburst after 0.05 s. (B) Illustrations of crystal growth observed for solutions of nfeAFP8 (a-d) and nfeAFP6 (e-h) at different temperatures. |
Ice crystal morphology
| PMID | 20853841 |
| DOI | 10.1021/bi100516e |
| Protein Name | SPnfe6 |
| Tag | GFP(N);Hexahistidine Tag(C). |
| Ice crystal morphology | Fig.6 Ice crystal morphology and growth in the presence of SPnfe6. Images of an ice crystal in 100 μM SPnfe6 at 0.05 ℃ of undercooling after 1 min (i) and 4 min (ii). The black bar represents a length of 50 μm. |
Ice crystal morphology
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6 |
| Tag | Hexahistidine Tag |
| Ice crystal morphology | Fig.2.A Morphological change in an ice crystal observed for 1 mM solutions. |
Thermal Hysteresis
| PMID | 30760774 |
| DOI | 10.1038/s41598-018-36546-2 |
| Protein Name | Zoarces elongatus AFPIII |
| Thermal Hysteresis | Fig.3.C Concentration dependence of thermal hysteresis evaluated for the five AFP samples. The A20L failed to halt the ice crystal growth, for which no TH activity was evaluated. |
Thermal Hysteresis
| PMID | 15654886 |
| DOI | 10.1111/j.1742-4658.2004.04490.x |
| Protein Name | nfeAFP6 |
| Thermal Hysteresis | Fig.6 TH activity measured using an osmometer (model OM 802; Vogel) as a function of concentration (mM) of type III AFP isoforms, nfeAFP2 (□), nfeAFP6 (◆), nfeAFP6△Lys (◇), nfeAFP8 (x), nfeAFP11 (○), nfeAFP13 in the absence of dithiothreitol (△), and nfeAFP13 in the presence of dithiothreitol (▲). The measurement was repeated three times using fresh samples, and mean values were plotted with error bars. |
Thermal Hysteresis
| PMID | 19187223 |
| DOI | 10.1111/j.1742-4658.2009.06887.x |
| Protein Name | nfeAFP6 (SP) |
| Thermal Hysteresis | Fig.4 (B) TH activities of nfeAFP6 (filled triangles), nfeAFP8 (filled circles) and a 1 : 1 mixture of the two isoforms (open circles) as a function of each total concentration. TH was evaluated as the difference between Tf and Tm (i.e. TH = |Tm-Tf|). For nfeAFP6, Tf = Tm. For nfeAFP8 and the 1 : 1 mixture, Tburst = Tf. (C) TH activity of nfeAFP6 in the presence of various proportions of nfeAFP8, with the total protein concentration adjusted to 0.1 mM. |
Thermal Hysteresis
| PMID | 19187223 |
| DOI | 10.1111/j.1742-4658.2009.06887.x |
| Protein Name | nfeAFP6 (SP) |
| Thermal Hysteresis | Fig.5 Concentration dependence of |Tm ) Tburst| for nfeAFP6 (open circles) and nfeAFP8 (filled circles). Tburst represents the temperature at which a burst of ice crystal growth occurs from the tip of the ice bipyramid. We were unable to define TH activity, but obtained a |Tm ) Tburst| value for nfeAFP6. For nfeAFP8, this value was identical to the TH activity (Fig. 4B). |
Thermal Hysteresis
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6 |
| Tag | Hexahistidine Tag |
| Thermal Hysteresis | Fig.2.B TH activities of wt and mutant AFPs as a function of each total concentration. TH is defined as the absolute value of the difference between Tm and Tburst. |
Fluorescence-based Ice Plane Affinity
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (wild type) |
| Fluorescence-based Ice Plane Affinity | Fig.1.C Definition of ice planes and FIPA patterns observed on a single ice crystal hemisphere. (C) FIPA pattern of a WT Zoarcidae-derived IBP containing A20. |
Fluorescence-based Ice Plane Affinity
| PMID | 30760774 |
| DOI | 10.1038/s41598-018-36546-2 |
| Protein Name | Zoarces elongatus AFPIII |
| Fluorescence-based Ice Plane Affinity | Fig.5.C Analysis of fluorescence-based ice plane affinity (FIPA) of AFPI-III, A20L, and Tis8 samples. |
Fluorescence-based Ice Plane Affinity
| PMID | 20853841 |
| DOI | 10.1021/bi100516e |
| Protein Name | SPnfe6 |
| Tag | GFP(N);Hexahistidine Tag(C). |
| Fluorescence-based Ice Plane Affinity | Fig.3 Binding of GFP-tagged SPnfe6 to an ice hemisphere. The hemisphere was mounted with its secondary prism plane normal to the coldfinger. Two distinct patches of fluorescence can be seen, with the lobes in each patch corresponding to pyramidal plane binding and the small patch of fluorescence linking each lobe being the signature for primary prism plane binding. |
Ice Recrystallization Inhibition
| PMID | 30760774 |
| DOI | 10.1038/s41598-018-36546-2 |
| Protein Name | Zoarces elongatus AFPIII |
| Ice Recrystallization Inhibition | Fig.2.B Photo examples of time-dependent changes in the ice grains in solvent (40% sucrose) and 1.5 μM solutions of AFPI, AFPII, AFPIII, A20L, and Tis8 at -6℃, taken from the last 20 min. The scale bars represent 50 μm. |
Ice Recrystallization Inhibition
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6 |
| Tag | Hexahistidine Tag |
| Ice Recrystallization Inhibition | Fig.3 Ice crystal images collected after complete freezing at 30 ℃ for 5 min (row 1) and after 30 min (row 2) and 4 h (row 3) at 6 ℃. |
Ice Binding Sites
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (wild type) |
| IBS | The Ice-Binding Site (IBS) contains protein residues T18,P19,A20,Q9,P12,N14,T15,A16,Q44. |
Ice Binding Sites
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6 |
| Tag | Hexahistidine Tag |
| IBS | The Ice-Binding Site (IBS) contains protein residues 9-21 and 41-44. |
Ice Grow Rate
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (wild type) |
| Ice Grow Rate | Fig.2 Dependence of the length of the a axis on the annealing time measured at the Tm -0.05 °C with 0.2 °C/min of cooling. The larger growth rate (μm/min) implies a weaker ability for the ice to inhibit growth. |
Ice Grow Rate
| PMID | 19187223 |
| DOI | 10.1111/j.1742-4658.2009.06887.x |
| Protein Name | nfeAFP6 (SP) |
| Ice Grow Rate | Tab.1 Ice growth rates determined for solutions of nfeAFP6, nfeAFP8 and the negative control (lysozyme) between Tm and Tburst. |
Ice Grow Rate
| PMID | 20853841 |
| DOI | 10.1021/bi100516e |
| Protein Name | SPnfe6 |
| Tag | GFP(N);Hexahistidine Tag(C). |
| Ice Grow Rate | Tab.1 Ice Crystal Growth Rates in the Presence of Various Type III AFP Iisoforms, Mutants, and Mixtures. |
Biological Cryoprotection Assessment
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6 |
| Tag | Hexahistidine Tag |
| Biological Cryoprotection Assessment | Fig.4 Beneficial effects of nfeAFPs on mouse ovarian tissue preservation upon vitrification and warming. Preservative effects were quantified in terms of on (A) intact follicle morphology and (B) ovarian follicle apoptosis. Follicle stability was assessed by (C) DNA DSBs, detected via cH2AX staining, and (D) DNA repair activity, measured by staining for Rad51. Different superscript letters indicate significant difference (P < 0.05). |
Ice plane
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (wild type) |
| Ice plane | The Ice-plane contains the pyramidal plane and the primary prism plane. |
Ice plane
| PMID | 19187223 |
| DOI | 10.1111/j.1742-4658.2009.06887.x |
| Protein Name | nfeAFP6 (SP) |
| Ice plane | The Ice-plane includes the pyramidal plane. |
Ice plane
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6 |
| Tag | Hexahistidine Tag |
| Ice plane | The Ice-plane includes pyramidal plane. |
Brief description
|
PMID: 29735675
DOI: 10.1073/pnas.1800635115 |
|
|
PMID: 30760774
DOI: 10.1038/s41598-018-36546-2 |
1. There is no obvious correlation between thermal hysteresis activity and ice recrystallization inhibition activity. For example, the thermal hysteresis values of AFP II and AFP III are similar, but their ice recrystallization inhibition efficiencies are significantly different.
2. Antifreeze proteins can change the morphology of ice crystals. Low concentrations of antifreeze proteins can trigger ice recrystallization inhibition activity, and high concentrations are not necessary. Excess antifreeze proteins may be harmful. |
|
PMID: 15654886
DOI: 10.1111/j.1742-4658.2004.04490.x |
1. There are multiple isoforms of type III antifreeze protein in Notched-fin eelpout, which can be divided into SP group, QAE1 group and QAE2 group.
2. The thermal hysteresis activities of different groups of antifreeze protein isoforms vary significantly. The nfeAFP8 in the QAE1 group has a relatively high thermal hysteresis activity, while the thermal hysteresis activities of nfeAFP2 and nfeAFP6 in the SP group are almost zero. 3. A small amount of "active" QAE1 isoforms (such as nfeAFP8) can significantly enhance the thermal hysteresis activity of "less active" SP isoforms (such as nfeAFP6), showing a cooperative effect. |
|
PMID: 19187223
DOI: 10.1111/j.1742-4658.2009.06887.x |
1. A minute amount of the active QAE isoform (nfeAFP8) confers thermal hysteresis activity on the SP isoform (nfeAFP6), which has no thermal hysteresis activity by itself.
2. The functions of nfeAFP6 and nfeAFP8 are equivalent with regard to the inhibition of growth from the tips of the ice bipyramid. |
|
PMID: 20853841
DOI: 10.1021/bi100516e |
1. SPnfe6 (SP isoform of type III AFP): It shares 55% identity with QAEop12, mainly binds the pyramidal plane, and has severely attenuated ability to bind the primary prism plane.
2. It lacks TH activity and cannot arrest ice growth, with a growth rate of 12.7 μm/min at 100 μM. It shapes ice into hexagonal bipyramids but allows steady growth of the bipyramids. |
|
PMID: 33598104
DOI: 10.1016/j.csbj.2021.01.016 |
1. Specific mutations in the QAE2 and SP isoforms can confer them with thermal hysteresis and ice recrystallization inhibition activities similar to those of the QAE1 isoform.
2. Factors such as the flexible structure of the first 310 helix and the distinct overall backbone conformation in inactive antifreeze proteins (QAE2 and SP isoforms) interfere with the hydrogen - bonding interaction with the primary prism plane of ice crystals, resulting in the loss of antifreeze activity. |
AFP013000001
| Mutation | A20L |
| Sequence |
Ice crystal morphology
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20L) |
| Ice crystal morphology | Fig.2.A Comparison of the strength of ice-growth inhibition between WT andmutant proteins. (A) Images of bipyramidal ice crystals captured for A20L, A20G,WT, A20T, A20V, and A20I mutants before and after 9 min of annealing time. Thearrow indicates the lengthof the a axis. |
Ice crystal morphology
| PMID | 30760774 |
| DOI | 10.1038/s41598-018-36546-2 |
| Protein Name | Zoarces elongatus AFPIII (A20L) |
| Ice crystal morphology | Fig.4.B Morphology of ice crystals observed for AFPI-III, A20L, and Tis8 at concentrations lower than their IRI endpoint. The hexagonal ice plate is created in the solutions of AFPI-AFPIII and A20L, while it is not facetted for Tis8 and similarly for the solvent (40% sucrose). |
Thermal Hysteresis
| PMID | 30760774 |
| DOI | 10.1038/s41598-018-36546-2 |
| Protein Name | Zoarces elongatus AFPIII (A20L) |
| Thermal Hysteresis | "For A20L, its growth inhibition ability was not enough to arrest ice growth, no TH value was evaluated." |
Fluorescence-based Ice Plane Affinity
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20L) |
| Fluorescence-based Ice Plane Affinity | Fig.1.E Definition of ice planes and FIPA patterns observed on a single ice crystal hemisphere. (E)Actual FIPA patterns observed for A20L, A20G, A20T, A20V, and A20I mutants.(Scale bars, 1 cm.) |
Fluorescence-based Ice Plane Affinity
| PMID | 30760774 |
| DOI | 10.1038/s41598-018-36546-2 |
| Protein Name | Zoarces elongatus AFPIII (A20L) |
| Fluorescence-based Ice Plane Affinity | Fig.5.C Analysis of fluorescence-based ice plane affinity (FIPA) of AFPI-III, A20L, and Tis8 samples. |
Ice Recrystallization Inhibition
| PMID | 30760774 |
| DOI | 10.1038/s41598-018-36546-2 |
| Protein Name | Zoarces elongatus AFPIII (A20L) |
| Ice Recrystallization Inhibition | Fig.2.B Photo examples of time-dependent changes in the ice grains in solvent (40% sucrose) and 1.5 μM solutions of AFPI, AFPII, AFPIII, A20L, and Tis8 at -6℃, taken from the last 20 min. The scale bars represent 50 μm. |
Ice Grow Rate
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20L) |
| Ice Grow Rate | Fig.2 Dependence of the length of the a axis on the annealing time measured at the Tm -0.05 °C with 0.2 °C/min of cooling. The larger growth rate (μm/min) implies a weaker ability for the ice to inhibit growth. |
Ice plane
| PMID | 30760774 |
| DOI | 10.1038/s41598-018-36546-2 |
| Protein Name | Zoarces elongatus AFPIII (A20L) |
| Ice plane | The Ice-plane includes the prism plane. |
Brief description
|
PMID: 29735675
DOI: 10.1073/pnas.1800635115 |
1. The A20I mutant has the strongest ice-binding ability, and the order of ice-growth inhibition strength is A20L < A20G < WT < A20T < A20V < A20I.
2. The location of the γ-CH3 group at the 20th residue plays a crucial role in the ice-binding ability of this IBP to the ice planes. |
|
PMID: 30760774
DOI: 10.1038/s41598-018-36546-2 |
1. There is no obvious correlation between thermal hysteresis activity and ice recrystallization inhibition activity. For example, the thermal hysteresis values of AFP II and AFP III are similar, but their ice recrystallization inhibition efficiencies are significantly different.
2. Antifreeze proteins can change the morphology of ice crystals. Low concentrations of antifreeze proteins can trigger ice recrystallization inhibition activity, and high concentrations are not necessary. Excess antifreeze proteins may be harmful. |
AFP013000002
| Mutation | A20G |
| Sequence |
Ice crystal morphology
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20G) |
| Ice crystal morphology | Fig.2.A Comparison of the strength of ice-growth inhibition between WT andmutant proteins. (A) Images of bipyramidal ice crystals captured for A20L, A20G,WT, A20T, A20V, and A20I mutants before and after 9 min of annealing time. Thearrow indicates the lengthof the a axis. |
Fluorescence-based Ice Plane Affinity
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20G) |
| Fluorescence-based Ice Plane Affinity | Fig.1.E Definition of ice planes and FIPA patterns observed on a single ice crystal hemisphere. (E)Actual FIPA patterns observed for A20L, A20G, A20T, A20V, and A20I mutants.(Scale bars, 1 cm.) |
Ice Grow Rate
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20G) |
| Ice Grow Rate | Fig.2 Dependence of the length of the a axis on the annealing time measured at the Tm -0.05 °C with 0.2 °C/min of cooling. The larger growth rate (μm/min) implies a weaker ability for the ice to inhibit growth. |
Brief description
|
PMID: 29735675
DOI: 10.1073/pnas.1800635115 |
1. The A20I mutant has the strongest ice-binding ability, and the order of ice-growth inhibition strength is A20L < A20G < WT < A20T < A20V < A20I.
2. The location of the γ-CH3 group at the 20th residue plays a crucial role in the ice-binding ability of this IBP to the ice planes. |
AFP013000003
| Mutation | A20T |
| Sequence |
Ice crystal morphology
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20T) |
| Ice crystal morphology | Fig.2.A Comparison of the strength of ice-growth inhibition between WT andmutant proteins. (A) Images of bipyramidal ice crystals captured for A20L, A20G,WT, A20T, A20V, and A20I mutants before and after 9 min of annealing time. Thearrow indicates the lengthof the a axis. |
Fluorescence-based Ice Plane Affinity
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20T) |
| Fluorescence-based Ice Plane Affinity | Fig.1.E Definition of ice planes and FIPA patterns observed on a single ice crystal hemisphere. (E)Actual FIPA patterns observed for A20L, A20G, A20T, A20V, and A20I mutants.(Scale bars, 1 cm.) |
Ice Grow Rate
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20T) |
| Ice Grow Rate | Fig.2 Dependence of the length of the a axis on the annealing time measured at the Tm -0.05 °C with 0.2 °C/min of cooling. The larger growth rate (μm/min) implies a weaker ability for the ice to inhibit growth. |
Brief description
|
PMID: 29735675
DOI: 10.1073/pnas.1800635115 |
1. The A20I mutant has the strongest ice-binding ability, and the order of ice-growth inhibition strength is A20L < A20G < WT < A20T < A20V < A20I.
2. The location of the γ-CH3 group at the 20th residue plays a crucial role in the ice-binding ability of this IBP to the ice planes. |
AFP013000004
| Mutation | A20V |
| Sequence |
Ice crystal morphology
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20V) |
| Ice crystal morphology | Fig.2.A Comparison of the strength of ice-growth inhibition between WT andmutant proteins. (A) Images of bipyramidal ice crystals captured for A20L, A20G,WT, A20T, A20V, and A20I mutants before and after 9 min of annealing time. Thearrow indicates the lengthof the a axis. |
Fluorescence-based Ice Plane Affinity
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20V) |
| Fluorescence-based Ice Plane Affinity | Fig.1.E Definition of ice planes and FIPA patterns observed on a single ice crystal hemisphere. (E)Actual FIPA patterns observed for A20L, A20G, A20T, A20V, and A20I mutants.(Scale bars, 1 cm.) |
Ice Grow Rate
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20V) |
| Ice Grow Rate | Fig.2 Dependence of the length of the a axis on the annealing time measured at the Tm -0.05 °C with 0.2 °C/min of cooling. The larger growth rate (μm/min) implies a weaker ability for the ice to inhibit growth. |
Brief description
|
PMID: 29735675
DOI: 10.1073/pnas.1800635115 |
1. The A20I mutant has the strongest ice-binding ability, and the order of ice-growth inhibition strength is A20L < A20G < WT < A20T < A20V < A20I.
2. The location of the γ-CH3 group at the 20th residue plays a crucial role in the ice-binding ability of this IBP to the ice planes. |
AFP013000005
| Mutation | A20I |
| Sequence |
Ice crystal morphology
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20I) |
| Ice crystal morphology | Fig.2.A Comparison of the strength of ice-growth inhibition between WT andmutant proteins. (A) Images of bipyramidal ice crystals captured for A20L, A20G,WT, A20T, A20V, and A20I mutants before and after 9 min of annealing time. Thearrow indicates the lengthof the a axis. |
Thermal Hysteresis
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20I) |
| Thermal Hysteresis | "Maximal thermal hysteresis activity for A20I was evaluated at ~1.2 °C." |
Fluorescence-based Ice Plane Affinity
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20I) |
| Fluorescence-based Ice Plane Affinity | Fig.1.E Definition of ice planes and FIPA patterns observed on a single ice crystal hemisphere. (E)Actual FIPA patterns observed for A20L, A20G, A20T, A20V, and A20I mutants.(Scale bars, 1 cm.) |
Ice Grow Rate
| PMID | 29735675 |
| DOI | 10.1073/pnas.1800635115 |
| Protein Name | NEF6 (A20I) |
| Ice Grow Rate | Fig.2 Dependence of the length of the a axis on the annealing time measured at the Tm -0.05 °C with 0.2 °C/min of cooling. The larger growth rate (μm/min) implies a weaker ability for the ice to inhibit growth. |
Brief description
|
PMID: 29735675
DOI: 10.1073/pnas.1800635115 |
1. The A20I mutant has the strongest ice-binding ability, and the order of ice-growth inhibition strength is A20L < A20G < WT < A20T < A20V < A20I.
2. The location of the γ-CH3 group at the 20th residue plays a crucial role in the ice-binding ability of this IBP to the ice planes. |
AFP013000006
| Mutation | 66del |
| Sequence |
Thermal Hysteresis
| PMID | 15654886 |
| DOI | 10.1111/j.1742-4658.2004.04490.x |
| Protein Name | nfe6△Lys |
| Thermal Hysteresis | Fig.6 TH activity measured using an osmometer (model OM 802; Vogel) as a function of concentration (mM) of type III AFP isoforms, nfeAFP2 (□), nfeAFP6 (◆), nfeAFP6△Lys (◇), nfeAFP8 (x), nfeAFP11 (○), nfeAFP13 in the absence of dithiothreitol (△), and nfeAFP13 in the presence of dithiothreitol (▲). The measurement was repeated three times using fresh samples, and mean values were plotted with error bars. |
Brief description
|
PMID: 15654886
DOI: 10.1111/j.1742-4658.2004.04490.x |
1. There are multiple isoforms of type III antifreeze protein in Notched-fin eelpout, which can be divided into SP group, QAE1 group and QAE2 group.
2. The thermal hysteresis activities of different groups of antifreeze protein isoforms vary significantly. The nfeAFP8 in the QAE1 group has a relatively high thermal hysteresis activity, while the thermal hysteresis activities of nfeAFP2 and nfeAFP6 in the SP group are almost zero. 3. A small amount of "active" QAE1 isoforms (such as nfeAFP8) can significantly enhance the thermal hysteresis activity of "less active" SP isoforms (such as nfeAFP6), showing a cooperative effect. |
AFP013000007
| Mutation | P19L,A20V |
| Sequence |
Ice crystal morphology
| PMID | 20853841 |
| DOI | 10.1021/bi100516e |
| Protein Name | SPnfe6_P19L/A20V |
| Tag | GFP(N);Hexahistidine Tag(C). |
| Ice crystal morphology | Fig.7.A Ice crystal morphology produced in a 100 μM solution of SPnfe6_P19L/A20V held 0.05 ℃ below the melting point after 1 min (i) and 15 min (ii). The black bar represents 50 μm. |
Thermal Hysteresis
| PMID | 20853841 |
| DOI | 10.1021/bi100516e |
| Protein Name | SPnfe6_P19L/A20V |
| Tag | GFP(N);Hexahistidine Tag(C). |
| Thermal Hysteresis | Fig.7.B Curve plotting the TH activity of SPnfe6_P19L/A20V (solid line) as a function of concentration. The TH activity of QAEop12 is also plotted for comparison (dashed line). |
Fluorescence-based Ice Plane Affinity
| PMID | 20853841 |
| DOI | 10.1021/bi100516e |
| Protein Name | SPnfe6_P19L/A20V |
| Tag | GFP(N);Hexahistidine Tag(C). |
| Fluorescence-based Ice Plane Affinity | Fig.5 Binding of GFP-tagged Spnfe6_P19L/A20V to an ice hemisphere. The hemisphere was mounted with its primary prism plane normal to the ice finger. |
Ice Grow Rate
| PMID | 20853841 |
| DOI | 10.1021/bi100516e |
| Protein Name | SPnfe6_P19L/A20V |
| Tag | GFP(N);Hexahistidine Tag(C). |
| Ice Grow Rate | Tab.1 Ice Crystal Growth Rates in the Presence of Various Type III AFP Iisoforms, Mutants, and Mixtures. |
Ice plane
| PMID | 20853841 |
| DOI | 10.1021/bi100516e |
| Protein Name | SPnfe6_P19L/A20V |
| Tag | GFP(N);Hexahistidine Tag(C). |
| Ice plane | The Ice-plane contains primary prism plane and the pyramidal plane. |
Brief description
|
PMID: 20853841
DOI: 10.1021/bi100516e |
1. The P19L/A20V mutations make it more QAE-like. It exhibits TH activity slightly higher than QAEop12 and significantly slows ice growth (0.4 μm/min at 100 μM).
2. GFP-tagged mutant shows broad fluorescence coverage on ice hemispheres, similar to QAEop12. Mixing 1% of this mutant with SPnfe6 drastically reduces the growth rate of SPnfe6. |
AFP013000008
| Mutation | P19L,A20V,G42S |
| Sequence |
Ice crystal morphology
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6_tri |
| Tag | Hexahistidine Tag |
| Ice crystal morphology | Fig.2.A Morphological change in an ice crystal observed for 1 mM solutions. |
Thermal Hysteresis
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6_tri |
| Tag | Hexahistidine Tag |
| Thermal Hysteresis | Fig.2.B TH activities of wt and mutant AFPs as a function of each total concentration. TH is defined as the absolute value of the difference between Tm and Tburst. |
Ice Recrystallization Inhibition
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6_tri |
| Tag | Hexahistidine Tag |
| Ice Recrystallization Inhibition | Fig.3 Ice crystal images collected after complete freezing at 30 ℃ for 5 min (row 1) and after 30 min (row 2) and 4 h (row 3) at 6 ℃. |
Biological Cryoprotection Assessment
| PMID | 33598104 |
| DOI | 10.1016/j.csbj.2021.01.016 |
| Protein Name | nfeAFP6_tri |
| Tag | Hexahistidine Tag |
| Biological Cryoprotection Assessment | Fig.4 Beneficial effects of nfeAFPs on mouse ovarian tissue preservation upon vitrification and warming. Preservative effects were quantified in terms of on (A) intact follicle morphology and (B) ovarian follicle apoptosis. Follicle stability was assessed by (C) DNA DSBs, detected via cH2AX staining, and (D) DNA repair activity, measured by staining for Rad51. Different superscript letters indicate significant difference (P < 0.05). |
Brief description
|
PMID: 33598104
DOI: 10.1016/j.csbj.2021.01.016 |
1. Specific mutations in the QAE2 and SP isoforms can confer them with thermal hysteresis and ice recrystallization inhibition activities similar to those of the QAE1 isoform.
2. Factors such as the flexible structure of the first 310 helix and the distinct overall backbone conformation in inactive antifreeze proteins (QAE2 and SP isoforms) interfere with the hydrogen - bonding interaction with the primary prism plane of ice crystals, resulting in the loss of antifreeze activity. |