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<big>'''β-Lactoglobulin'''</big>
----

<Structure load='1beb' size='400' frame='true' align='right' caption='The dimer interface of β-lactoglobulin lattice X' scene='Insert optional scene name here' />

== Introduction ==

β-Lactoglobulin (β-LG)is the major whey protein of cow’s milk with a concentration of 0.3 g/100 mL <ref>PMID:14254409</ref> and was first isolated in 1934 <ref>http://www.jbc.org/content/104/2/359.citation</ref>. It is a globular protein consisting of 162 AA with a relative molecular mass of 18.4 kDa.

Its amino-acid sequence and 3-dimensional structure show that it is a lipocalin, a widely diverse family, most of which bind small hydrophobic ligands and thus may act as specific transporters, as does serum retinol binding protein <ref>PMID:15259212</ref>.

Under physiological conditions beta-lactoglobulin exists as an equilibrium mixture of monomeric and dimeric forms.
Subcellular location: Secreted.
Tissue specificity: Synthesized in mammary gland and secreted in milk.
Post-translational modification : Alternate disulfide bonds occur in equal amounts in all variants examined.
Allergenic properties:Causes an allergic reaction in human. Is one of the causes of cow's milk allergy.

Bovine b-lactoglobulin (β-Lg) is a much studied and commercially important whey protein with an as yet undetermined function,although it is of obvious nutritional value. b-Lg binds a variety of ligands and by comparison of the general structures of these
molecules together with several competition studies, it appears that there are at least 3 independent binding sites. In the absence of direct crystallographic evidence, a preliminary modelling study reveals that there is an internal cavity which can readily accommodate retinol in a manner similar to the related lipocalin, retinol-binding protein. On the outer surface, a solvent-accessible hydrophobic cleft runs between the 3-turn a-helix that is packed against the outer surface of the b-barrel. This cleft can accommodate fatty acids like palmitate and stearate.
<ref>http://www.sciencedirect.com/science/article/pii/S0958694698000211</ref>

β-Lactoglobulin is a small protein, soluble in dilute salt solution as befits a globulin, with 162 amino acid residues (Mr ∼18,400) that fold up into an 8-stranded, antiparallel β-barrel with a 3-turn α-helix on the outer surface and a ninth β-strand flanking the first strand.

Function:Primary component of whey, it binds retinol and is probably involved in the transport of that molecule.<ref>PMID:15259212</ref>.

[[Image:Beta_lac_2Q2M.png|thumb|right|320px|Bovine Beta-Lactoglobulin Native]]

===Background ===
class of protein :Belongs to the calycin superfamily. Lipocalin family.
overall function of Lipocalin family: The lipocalins are a family of proteins which transport small hydrophobic molecules such as steroids, bilins, retinoids, and lipids.
Lipocalins have been associated with many biological processes, among them immune response, pheromone transport, biological prostaglandin synthesis, retinoid binding, and cancer cell interactions.

organisms:These proteins are found in gram negative bacteria, vertebrate cells, and invertebrate cells, and in plants.

===Lipocalin Proteins===
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>

The lipocalin family is a large and diverse family of proteins with functions varying from insect camouflage to small hydrophobic molecule transport typified by the serum retinol-binding protein <ref>http://www.biochemj.org/bj/318/bj3180001.htm</ref> The crystal structures so far determined reveal the typical lipocalin to be an eight-stranded antiparallel β-barrel arranged to form a conical central calyx or cavity in which the hydrophobic ligand is located.<ref>PMID:6540172</ref>
<ref>PMID:9867826</ref>

== Structure of β-LG ==

<Structure load='1beb' size='400' frame='true' align='right' caption='β-Lactoglobulin (β-LG) (variant A)' scene='Insert optional scene name here' />

βLG consists of 162 amino acid residues (18 kDa), containing two disulfide bonds (Cys 66–Cys 160 and Cys 106–Cys 119) and a free thiol (Cys 121). Structures of βLG have been reported by several groups with X-ray crystallography [19–21] and solution NMR [29,40,41]. It is a predominantly β-sheet protein. The β-barrel, or so called calyx, is conical and is made of two β-sheets: the B–D strands and N-terminal half of the A strand (denoted AN) form one sheet, and the E–H strands and C-terminal half of the A strand (denoted AC)form the other. On the outer surface of the β-barrel, between the G and H strands, is the 3-turn α-helix. The loops that connect the β-strands at the closed end of the calyx, BC, DE, and FG,are generally quite short, whereas those at the open end, AB, CD, EF,and GH, are significantly longer and more flexible [19]. In the calyx,there is a large central cavity which is surrounded by hydrophobic residues and is accessible to solvent. This cavity provides the principal ligand-binding site. βLG contains two tryptophan residues,Trp 19 on the A strand and Trp 61 on the C strand. The former is buried in the hydrophobic core whereas the latter is exposed to the solvent in the native structure, making them useful probes for monitoring site-specific conformational changes. In addition, studies on the monomer–dimer equilibrium [30,32,42,43] and the reactivity of the thiol group of Cys121 deeply buried between the α-helix and H strand [44–48] revealed other important properties of βLG.<ref>PMID:19362581</ref> the stability of the structure should depend so heavily upon the external loop around residue 64 or the beta strand with the free thiol<ref>PMID:9115437</ref>

At physiological conditions, bovine b-lactoglobulin forms a dimer, with each monomer consisting of 162 amino acid residues and characterized by a molecular mass of 18,350. Below pH 3, the dimer dissociates into <scene name='56/564055/Native_beta-lactoglobulin/1'>monomers</scene> which preserve their native conformation. Genetically, b-lactoglobulin may exist as one of several variants, among which the variants A and B are the most abundant. The A and B variants of the protein differ from each other by amino acid residues at positions Asp64 (Gly64 in variant B) and Val118 (Ala118 in variant B). These differences in primary structure render the two variants slightly different with respect to isoelectric point, solubility, self-association properties, as well as pressure and temperature stability. However, the structural characteristics of the A and B variants of bovine b-lactoglobulin are virtually indistinguishable. In its native state, b-lactoglobulin is a predominantly b-sheet protein containing nine b-strands and three a-helices. The core of the protein is formed by a flattened b-barrel (a calyx) composed of eight antiparallel b-strands (A to H).<ref>PMID:11734004</ref>

Ligands
Most lipocalins bind small hydrophobic molecules within the central cup or calyx. The true function of b-Lg is unknown, but it has been suggested that it is involved in
the transport of retinol and/or fatty acids [8,50]. It binds retinol with a higher affinity than does RBP [51] and, as with RBP, specific binding of retinol to b-Lg has been observed in the small intestine of the neonatal calf [3]. The structure of RBP with retinol bound within the hydrophobic calyx has been solved [2] and retinol was successfully modelled into our previous b-Lg structure [3].b-Lg contains two tryptophans, Trp19 and Trp61, and their fluorescence is altered when retinol is bound [51].<ref>PMID:9115437</ref>

==Molecular mechanism of the Tanford transition==
Above pH 6.5, b-lactoglobulin undergoes the so-called Tanford transition which is triggered by protonation of Glu89 exhibiting an anomalously
high pKa value. The Tanford transition involves displacement of the loop EF (residues 85 to 90) that acts as a lid which closes the protein interior/binding site below pH 7.3 and opens it at higher pH. The Tanford transition may involve some other structural changes as well. For example, the transition is accompanied by a change in the microenvironment of Tyr428 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. It should be noted that all transitions that take place between pH 2 and pH 9 do not cause any appreciable changes in the nativelike b-barrel conformation of b-lactoglobulin.
<ref>PMID:11734004</ref>

The structures of the trigonal crystal form of bovine beta-lactoglobulin variant A at pH 6.2, 7.1, and 8.2 have been determined by X-ray diffraction methods at a resolution of 2.56, 2. 24, and 2.49 A, respectively. 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 85-90, 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>

Dimeric Lactoglobulin molecules exist in the open conformation at basic pH, whereas they exist in the closed conformation at acidic pH, after undergoing Tanford transition around neutral pH.<ref>PMID:17932936</ref>

The Tanford transition is a conformational change of bovine beta-lactoglobulin (betaLG) 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 is responsible for the Tanford transition, with the loop closing the hydrophobic cavity of the beta-barrel of the betaLG molecule below pH 7 and flipping to open the cavity above pH 7.<ref>PMID:16368109</ref>

==Implications or possible application==

Retinol and Palmitate Binding
Ever since the fortuitous observation that beta-lactoglobulin (beta-Lg), the major whey protein in the milk of ruminants, bound retinol, the details of the binding have been controversial. beta-Lg is a lipocalin, like plasma retinol-binding protein, so that ligand association was expected to make use of the central cavity in the protein.<ref>PMID:12054801</ref>
A cocrystallized β-Lg with palmitic acid, and the refined structure (R = 0.204, R free = 0.240 for 6,888 reflections to 2.5-Å resolution) 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.<ref>PMID:9867826</ref>

Antioxidant Nature
Cross-linking the free thiol groups of beta-LG by heating (100 degrees C for 2 min), or chemically modifying the beta-LG by carboxymethylation to block the thiol groups resulted in a substantial loss of antioxidant activity. The data suggest that Cys-121 plays an essential role in the antioxidant nature of beta-LG.<ref>PMID:17235131</ref>

==Other β-Lactoglobulin 3D Structures and complexes==

[[1b8e]] - Crystal structure of the Bovine beta-Lactoglobulin (Isoforms A and B) in orthorombic space group

[[1qg5]] - Crystal structure of the Bovine beta-Lactoglobulin (Isoforms A)

[[1beb]] - Bovine beta-Lactoglobulin, Lattice X

[[1cj5]] - Bovine beta-Lactoglobulin A

[[1gx8]] - Bovine beta-Lactoglobulin complexed with Retinol, Trigonal Lattice Z

[[1gx9]] - Bovine beta-Lactoglobulin complexed with Retinoic acid, Trigonal Lattice Z

[[1gxa]] - Bovine beta-Lactoglobulin complexed with Retinol and Palmitic acid, Trigonal Lattice Z

[[1b0o]] - Bovine beta-Lactoglobulin complexed with Palmitate, Lattice Z

[[1bsy]] [[2blg]] [[3blg]] - Structural Basis of the Tanford Transitioon of Bovine beta-Lactoglobulin from crystal structures at 3 pH values

== References ==
<references/>

== Proteopedia Page Contributors and Editors ==
[[User:Rini Triani|Rini Triani]]

[[Category:Bos taurus]]
[[Category:Lipocalin]]

Sandbox Reserved 779

2013-12-04T09:35:58Z

Rini Triani:

[[Image:Example.jpg]
{{User:Michael_B._Goshe/Template_BCH455_555}}


<big>'''β-Lactoglobulin'''</big>
----

<Structure load='1beb' size='400' frame='true' align='right' caption='The dimer interface of β-lactoglobulin lattice X' scene='Insert optional scene name here' />

== Introduction ==

β-Lactoglobulin (β-LG)is the major whey protein of cow’s milk with a concentration of 0.3 g/100 mL <ref>PMID:14254409</ref> and was first isolated in 1934 <ref>http://www.jbc.org/content/104/2/359.citation</ref>. It is a globular protein consisting of 162 AA with a relative molecular mass of 18.4 kDa.

Its amino-acid sequence and 3-dimensional structure show that it is a lipocalin, a widely diverse family, most of which bind small hydrophobic ligands and thus may act as specific transporters, as does serum retinol binding protein <ref>PMID:15259212</ref>.

Under physiological conditions beta-lactoglobulin exists as an equilibrium mixture of monomeric and dimeric forms.
Subcellular location: Secreted.
Tissue specificity: Synthesized in mammary gland and secreted in milk.
Post-translational modification : Alternate disulfide bonds occur in equal amounts in all variants examined.
Allergenic properties:Causes an allergic reaction in human. Is one of the causes of cow's milk allergy.

Bovine b-lactoglobulin (β-Lg) is a much studied and commercially important whey protein with an as yet undetermined function,although it is of obvious nutritional value. b-Lg binds a variety of ligands and by comparison of the general structures of these
molecules together with several competition studies, it appears that there are at least 3 independent binding sites. In the absence of direct crystallographic evidence, a preliminary modelling study reveals that there is an internal cavity which can readily accommodate retinol in a manner similar to the related lipocalin, retinol-binding protein. On the outer surface, a solvent-accessible hydrophobic cleft runs between the 3-turn a-helix that is packed against the outer surface of the b-barrel. This cleft can accommodate fatty acids like palmitate and stearate.
<ref>http://www.sciencedirect.com/science/article/pii/S0958694698000211</ref>

β-Lactoglobulin is a small protein, soluble in dilute salt solution as befits a globulin, with 162 amino acid residues (Mr ∼18,400) that fold up into an 8-stranded, antiparallel β-barrel with a 3-turn α-helix on the outer surface and a ninth β-strand flanking the first strand.

Function:Primary component of whey, it binds retinol and is probably involved in the transport of that molecule.<ref>PMID:15259212</ref>.

[[Image:Beta_lac_2Q2M.png|thumb|right|320px|Bovine Beta-Lactoglobulin Native]]

===Background ===
class of protein :Belongs to the calycin superfamily. Lipocalin family.
overall function of Lipocalin family: The lipocalins are a family of proteins which transport small hydrophobic molecules such as steroids, bilins, retinoids, and lipids.
Lipocalins have been associated with many biological processes, among them immune response, pheromone transport, biological prostaglandin synthesis, retinoid binding, and cancer cell interactions.

organisms:These proteins are found in gram negative bacteria, vertebrate cells, and invertebrate cells, and in plants.

===Lipocalin Proteins===
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>

The lipocalin family is a large and diverse family of proteins with functions varying from insect camouflage to small hydrophobic molecule transport typified by the serum retinol-binding protein <ref>http://www.biochemj.org/bj/318/bj3180001.htm</ref> The crystal structures so far determined reveal the typical lipocalin to be an eight-stranded antiparallel β-barrel arranged to form a conical central calyx or cavity in which the hydrophobic ligand is located.<ref>PMID:6540172</ref>
<ref>PMID:9867826</ref>

== Structure of β-LG ==

<Structure load='1beb' size='400' frame='true' align='right' caption='β-Lactoglobulin (β-LG) (variant A)' scene='Insert optional scene name here' />

βLG consists of 162 amino acid residues (18 kDa), containing two disulfide bonds (Cys 66–Cys 160 and Cys 106–Cys 119) and a free thiol (Cys 121). Structures of βLG have been reported by several groups with X-ray crystallography [19–21] and solution NMR [29,40,41]. It is a predominantly β-sheet protein. The β-barrel, or so called calyx, is conical and is made of two β-sheets: the B–D strands and N-terminal half of the A strand (denoted AN) form one sheet, and the E–H strands and C-terminal half of the A strand (denoted AC)form the other. On the outer surface of the β-barrel, between the G and H strands, is the 3-turn α-helix. The loops that connect the β-strands at the closed end of the calyx, BC, DE, and FG,are generally quite short, whereas those at the open end, AB, CD, EF,and GH, are significantly longer and more flexible [19]. In the calyx,there is a large central cavity which is surrounded by hydrophobic residues and is accessible to solvent. This cavity provides the principal ligand-binding site. βLG contains two tryptophan residues,Trp 19 on the A strand and Trp 61 on the C strand. The former is buried in the hydrophobic core whereas the latter is exposed to the solvent in the native structure, making them useful probes for monitoring site-specific conformational changes. In addition, studies on the monomer–dimer equilibrium [30,32,42,43] and the reactivity of the thiol group of Cys121 deeply buried between the α-helix and H strand [44–48] revealed other important properties of βLG.<ref>PMID:19362581</ref> the stability of the structure should depend so heavily upon the external loop around residue 64 or the beta strand with the free thiol<ref>PMID:9115437</ref>

At physiological conditions, bovine b-lactoglobulin forms a dimer, with each monomer consisting of 162 amino acid residues and characterized by a molecular mass of 18,350. Below pH 3, the dimer dissociates into <scene name='56/564055/Native_beta-lactoglobulin/1'>monomers</scene> which preserve their native conformation. Genetically, b-lactoglobulin may exist as one of several variants, among which the variants A and B are the most abundant. The A and B variants of the protein differ from each other by amino acid residues at positions Asp64 (Gly64 in variant B) and Val118 (Ala118 in variant B). These differences in primary structure render the two variants slightly different with respect to isoelectric point, solubility, self-association properties, as well as pressure and temperature stability. However, the structural characteristics of the A and B variants of bovine b-lactoglobulin are virtually indistinguishable. In its native state, b-lactoglobulin is a predominantly b-sheet protein containing nine b-strands and three a-helices. The core of the protein is formed by a flattened b-barrel (a calyx) composed of eight antiparallel b-strands (A to H).<ref>PMID:11734004</ref>

Ligands
Most lipocalins bind small hydrophobic molecules within the central cup or calyx. The true function of b-Lg is unknown, but it has been suggested that it is involved in
the transport of retinol and/or fatty acids [8,50]. It binds retinol with a higher affinity than does RBP [51] and, as with RBP, specific binding of retinol to b-Lg has been observed in the small intestine of the neonatal calf [3]. The structure of RBP with retinol bound within the hydrophobic calyx has been solved [2] and retinol was successfully modelled into our previous b-Lg structure [3].b-Lg contains two tryptophans, Trp19 and Trp61, and their fluorescence is altered when retinol is bound [51].<ref>PMID:9115437</ref>

==Mechanism of action==

===Molecular mechanism of the Tanford transition===
Above pH 6.5, b-lactoglobulin undergoes the so-called Tanford transition which is triggered by protonation of Glu89 exhibiting an anomalously
high pKa value. The Tanford transition involves displacement of the loop EF (residues 85 to 90) that acts as a lid which closes the protein interior/binding site below pH 7.3 and opens it at higher pH. The Tanford transition may involve some other structural changes as well. For example, the transition is accompanied by a change in the microenvironment of Tyr428 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. It should be noted that all transitions that take place between pH 2 and pH 9 do not cause any appreciable changes in the nativelike b-barrel conformation of b-lactoglobulin.
<ref>PMID:11734004</ref>

The structures of the trigonal crystal form of bovine beta-lactoglobulin variant A at pH 6.2, 7.1, and 8.2 have been determined by X-ray diffraction methods at a resolution of 2.56, 2. 24, and 2.49 A, respectively. 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 85-90, 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>

Dimeric Lactoglobulin molecules exist in the open conformation at basic pH, whereas they exist in the closed conformation at acidic pH, after undergoing Tanford transition around neutral pH.<ref>PMID:17932936</ref>

The Tanford transition is a conformational change of bovine beta-lactoglobulin (betaLG) 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 is responsible for the Tanford transition, with the loop closing the hydrophobic cavity of the beta-barrel of the betaLG molecule below pH 7 and flipping to open the cavity above pH 7.<ref>PMID:16368109</ref>

==Implications or possible application==

Retinol Binding
Ever since the fortuitous observation that beta-lactoglobulin (beta-Lg), the major whey protein in the milk of ruminants, bound retinol, the details of the binding have been controversial. beta-Lg is a lipocalin, like plasma retinol-binding protein, so that ligand association was expected to make use of the central cavity in the protein.<ref>PMID:12054801</ref>

Antioxidant Nature
Cross-linking the free thiol groups of beta-LG by heating (100 degrees C for 2 min), or chemically modifying the beta-LG by carboxymethylation to block the thiol groups resulted in a substantial loss of antioxidant activity. The data suggest that Cys-121 plays an essential role in the antioxidant nature of beta-LG.<ref>PMID:17235131</ref>

==Other β-Lactoglobulin 3D Structures and complexes==

[[1b8e]] - Crystal structure of the Bovine beta-Lactoglobulin (Isoforms A and B) in orthorombic space group

[[1qg5]] - Crystal structure of the Bovine beta-Lactoglobulin (Isoforms A)

[[1beb]] - Bovine beta-Lactoglobulin, Lattice X

[[1cj5]] - Bovine beta-Lactoglobulin A

[[1gx8]] - Bovine beta-Lactoglobulin complexed with Retinol, Trigonal Lattice Z

[[1gx9]] - Bovine beta-Lactoglobulin complexed with Retinoic acid, Trigonal Lattice Z

[[1gxa]] - Bovine beta-Lactoglobulin complexed with Retinol and Palmitic acid, Trigonal Lattice Z

[[1b0o]] - Bovine beta-Lactoglobulin complexed with Palmitate, Lattice Z

[[1bsy]] [[2blg]] [[3blg]] - Structural Basis of the Tanford Transitioon of Bovine beta-Lactoglobulin from crystal structures at 3 pH values

== References ==
<references/>

== Proteopedia Page Contributors and Editors ==
[[User:Rini Triani|Rini Triani]]

[[Category:Bos taurus]]
[[Category:Lipocalin]]