Sandbox Reserved 779: Difference between revisions

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===Lipocalin Proteins===

β-Lactoglobulin belongs to the calycin superfamily and Lipocalin family. Lipocalins are typically small (160-180 residues in length), extracellular proteins sharing several common molecular recognition properties: the binding of small, principally hydrophobic molecules (such as retinol); binding to specific cell-surface receptors; and the formation of covalent and non-covalent complexes with other soluble macromolecules. Although they have been classified mainly as transport proteins

β-Lactoglobulin belongs to the calycin superfamily and Lipocalin family. Lipocalins are typically small (160-180 residues in length), extracellular proteins sharing several common molecular recognition properties: the binding of small, principally hydrophobic molecules (such as retinol); binding to specific cell-surface receptors; and the formation of covalent and non-covalent complexes with other soluble macromolecules. Although they have been classified mainly as transport proteins.

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===Dimer/Monomer===

At physiological conditions, majority of bovine b-lactoglobulin forms a dimer (Fig. 3). Below pH 3, the dimer dissociates into monomers ~~(Fig. 4)~~ which maintain their native conformation.

At physiological conditions, majority of bovine b-lactoglobulin forms a dimer (Fig. 3). Below pH 3, the dimer dissociates into monomers which maintain their native conformation.

[[Image:BLG_Dimer_1BEB_Chain_A&B.png|thumb|right|240px|Figure 3. Bovine β-Lactoglobulin Dimer_[[1BEB]]]]

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Co-crystallized β-Lg with palmitic acid (3D Model 2._1B0O) , and the refined structure reveals that the ligand binds in the central cavity in a manner similar to the binding of retinol to the related lipocalin, serum retinol-binding protein. The carboxyl group binds to both Lys-60 and Lys-69 at the entrance to the cavity. The hydrophobic tail stretches in an almost fully extended conformation into the center of the protein.<ref>PMID:9867826</ref>

In addition, studies on the monomer–dimer equilibrium and the reactivity of the thiol group of Cys121 which deeply buried between the α-helix and H strand revealed other important properties of β-LG. The stability of the structure also depend so heavily upon the external loop around residue 64 or the β strand with the free thiol.<ref>PMID:9115437</ref>

[[Image:Vitamin D3 binding to the beta-lac calyx and dimer interface.jpg|thumb|left|240px|Figure 4. Vitamin D3 binding to the beta-lac calyx and dimer interface. The BLG dimer is shown with the four experimentally determined VD3 ligands at their respective binding sites. The arrowheads indicate entrance to the calyx. (Dominguez-Ramirez et al, 2013),ref.PMID:24255705]]

β-Lg has binding sites for hydrophobic ligands like fatty acids, [[retinoic acid]], and Vitamin D3 ([[cholecalciferol]]) and [[lactose]] [[ligands]] <ref>PMID:24255705</ref> For hydrophobic ligands two sites have been postulated, one inside the calyx and the other at the dimer interface, on the outer surface of the protein between the α-helix and the β-barrel. The accessibility to the calyx is pH-dependent.

β-Lg has binding sites for hydrophobic ligands like fatty acids, retinoic acid, and Vitamin D3 (cholecalciferol) and lactose [[ligands]] <ref>PMID:24255705</ref> For hydrophobic ligands two sites have been postulated, one inside the calyx and the other at the dimer interface, on the outer surface of the protein between the α-helix and the β-barrel (Fig. 4). The accessibility to the calyx is pH-dependent.

NMR and Xray analysis showed that the access is mediated by the mobile EF loop. All the structures with ligands bound to the calyx exhibit an open EF loop, suggesting that this site is accessible at neutral pH.

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=='''Molecular mechanism of the Tanford transition'''==

The Tanford transition is a conformational change of bovine β-lactoglobulin occurring at around pH 7, identified originally on the basis of optical rotatory dispersion and the accessibility of a thiol group. X-ray analysis has suggested that a conformational change to the EF-loop (residues I84 -N90) (Fig. 4) is responsible for the Tanford transition, with the loop closing the hydrophobic cavity of the β-barrel of the β-LG molecule below pH 7 and flipping to open the cavity above pH 7.<ref>PMID:16368109</ref>Tanford transition is triggered by protonation of Glu89 exhibiting an anomalously high pKa value.

The Tanford transition is a conformational change of bovine β-lactoglobulin occurring at around pH 7, identified originally on the basis of optical rotatory dispersion and the accessibility of a thiol group. X-ray analysis has suggested that a conformational change to the EF-loop (residues I84 -N90) (Fig. 5) is responsible for the Tanford transition, with the loop closing the hydrophobic cavity of the β-barrel of the β-LG molecule below pH 7 and flipping to open the cavity above pH 7.<ref>PMID:16368109</ref>Tanford transition is triggered by protonation of Glu89 exhibiting an anomalously high pKa value.

The Tanford transition may involve some other structural changes as well. For example, the transition is accompanied by a change in the microenvironment of Tyr42 and causes an alteration in the relative orientation of monomers in the dimer by as much as 5 degrees, which breaks a number of intersubunit hydrogen bonds. <ref>PMID:11734004</ref>

[[Image:BLG_monomer_showing_EF_Loop.png|thumb|left|240px|Figure 4. Bovine β-Lactoglobulin Monomer showing EF loop (colored yellow)]]

[[Image:BLG_monomer_showing_EF_Loop.png|thumb|left|240px|Figure 5. Bovine β-Lactoglobulin Monomer showing EF loop (colored yellow)]]

The structures of the trigonal crystal form of bovine β-lactoglobulin variant A at pH 6.2, 7.1, and 8.2 have been determined by X-ray diffraction methods. The glutamate side chain of residue 89 is buried at pH 6.2 and becomes exposed at pH 7.1 and 8.2. This conformational change, involving the loop 84-90, provides a structural basis for a variety of pH-dependent chemical, physical, and spectroscopic phenomena, collectively known as the Tanford transition.<ref>PMID:9760236</ref>It was found that the stereochemical environment of Tyr42 changes significantly with pH variation between pH 7 and pH 8. This may provide a structural explanation for an as yet unexplained feature of the Tanford transition, namely the increase in exposure of a tyrosine residue.<ref>PMID:11168385</ref>

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@@ Line 14: / Line 14: @@
 ===Lipocalin Proteins===
-β-Lactoglobulin belongs to the calycin superfamily and Lipocalin family. Lipocalins are typically small (160-180 residues in length), extracellular proteins sharing several common molecular recognition properties: the binding of small, principally hydrophobic molecules (such as retinol); binding to specific cell-surface receptors; and the formation of covalent and non-covalent complexes with other soluble macromolecules. Although they have been classified mainly as transport proteins
+β-Lactoglobulin belongs to the calycin superfamily and Lipocalin family. Lipocalins are typically small (160-180 residues in length), extracellular proteins sharing several common molecular recognition properties: the binding of small, principally hydrophobic molecules (such as retinol); binding to specific cell-surface receptors; and the formation of covalent and non-covalent complexes with other soluble macromolecules. Although they have been classified mainly as transport proteins.
 <ref>PMID:11058743</ref>
@@ Line 30: / Line 30: @@
 ===Dimer/Monomer===
-At physiological conditions, majority of bovine b-lactoglobulin forms a dimer (Fig. 3). Below pH 3, the dimer dissociates into monomers (Fig. 4) which maintain their native conformation.
+At physiological conditions, majority of bovine b-lactoglobulin forms a dimer (Fig. 3). Below pH 3, the dimer dissociates into monomers which maintain their native conformation.
 [[Image:BLG_Dimer_1BEB_Chain_A&B.png|thumb|right|240px|Figure 3. Bovine β-Lactoglobulin Dimer_[[1BEB]]]]
@@ Line 42: / Line 42: @@
 Co-crystallized β-Lg with palmitic acid (3D Model 2._1B0O) , and the refined structure reveals that the ligand binds in the central cavity in a manner similar to the binding of retinol to the related lipocalin, serum retinol-binding protein. The carboxyl group binds to both Lys-60 and Lys-69 at the entrance to the cavity. The hydrophobic tail stretches in an almost fully extended conformation into the center of the protein.<ref>PMID:9867826</ref>
 In addition, studies on the monomer–dimer equilibrium and the reactivity of the thiol group of Cys121 which deeply buried between the α-helix and H strand revealed other important properties of β-LG. The stability of the structure also depend so heavily upon the external loop around residue 64 or the β strand with the free thiol.<ref>PMID:9115437</ref>
+[[Image:Vitamin D3 binding to the beta-lac calyx and dimer interface.jpg|thumb|left|240px|Figure 4. Vitamin D3 binding to the beta-lac calyx and dimer interface. The BLG dimer is shown with the four experimentally determined VD3 ligands at their respective binding sites. The arrowheads indicate entrance to the calyx. (Dominguez-Ramirez et al, 2013),ref.PMID:24255705]]
-β-Lg has binding sites for hydrophobic ligands like fatty acids, [[retinoic acid]], and Vitamin D3 ([[cholecalciferol]]) and [[lactose]] [[ligands]] <ref>PMID:24255705</ref> For hydrophobic ligands two sites have been postulated, one inside the calyx and the other at the dimer interface, on the outer surface of the protein between the α-helix and the β-barrel.  The accessibility to the calyx is pH-dependent.
+β-Lg has binding sites for hydrophobic ligands like fatty acids, retinoic acid, and Vitamin D3 (cholecalciferol) and lactose [[ligands]] <ref>PMID:24255705</ref> For hydrophobic ligands two sites have been postulated, one inside the calyx and the other at the dimer interface, on the outer surface of the protein between the α-helix and the β-barrel (Fig. 4).  The accessibility to the calyx is pH-dependent.
 NMR and Xray analysis showed that the access is mediated by the mobile EF loop. All the structures with ligands bound to the calyx exhibit an open EF loop, suggesting that this site is accessible at neutral pH.
 <Structure load='1b0o' size='300' frame='true' align='right' caption='3D model 2._β-lactoglobulin complexed with Palmitate, lattice Z_1B0O' scene='Insert optional scene name here' />
@@ Line 52: / Line 52: @@
 =='''Molecular mechanism of the Tanford transition'''==
-The Tanford transition is a conformational change of bovine β-lactoglobulin occurring at around pH 7, identified originally on the basis of optical rotatory dispersion and the accessibility of a thiol group. X-ray analysis has suggested that a conformational change to the EF-loop (residues I84 -N90) (Fig. 4) is responsible for the Tanford transition, with the loop closing the hydrophobic cavity of the β-barrel of the β-LG molecule below pH 7 and flipping to open the cavity above pH 7.<ref>PMID:16368109</ref>Tanford transition is triggered by protonation of Glu89 exhibiting an anomalously high pKa value.
+The Tanford transition is a conformational change of bovine β-lactoglobulin occurring at around pH 7, identified originally on the basis of optical rotatory dispersion and the accessibility of a thiol group. X-ray analysis has suggested that a conformational change to the EF-loop (residues I84 -N90) (Fig. 5) is responsible for the Tanford transition, with the loop closing the hydrophobic cavity of the β-barrel of the β-LG molecule below pH 7 and flipping to open the cavity above pH 7.<ref>PMID:16368109</ref>Tanford transition is triggered by protonation of Glu89 exhibiting an anomalously high pKa value.
 The Tanford transition may involve some other structural changes as well. For example, the transition is accompanied by a change in the microenvironment of Tyr42 and causes an alteration in the relative orientation of monomers in the dimer by as much as 5 degrees, which breaks a number of intersubunit hydrogen bonds. <ref>PMID:11734004</ref>
-[[Image:BLG_monomer_showing_EF_Loop.png|thumb|left|240px|Figure 4. Bovine β-Lactoglobulin Monomer showing EF loop (colored yellow)]]
+[[Image:BLG_monomer_showing_EF_Loop.png|thumb|left|240px|Figure 5. Bovine β-Lactoglobulin Monomer showing EF loop (colored yellow)]]
 The structures of the trigonal crystal form of bovine β-lactoglobulin variant A at pH 6.2, 7.1, and 8.2 have been determined by X-ray diffraction methods. The glutamate side chain of residue 89 is buried at pH 6.2 and becomes exposed at pH 7.1 and 8.2. This conformational change, involving the loop 84-90, provides a structural basis for a variety of pH-dependent chemical, physical, and spectroscopic phenomena, collectively known as the Tanford transition.<ref>PMID:9760236</ref>It was found that the stereochemical environment of Tyr42 changes significantly with pH variation between pH 7 and pH 8. This may provide a structural explanation for an as yet unexplained feature of the Tanford transition, namely the increase in exposure of a tyrosine residue.<ref>PMID:11168385</ref>