Mlk og molekylrbiologi

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Mælk og molekylærbiologi
Torben Ellebæk Petersen Protein Chemistry
Laboratory Department of Molecular Biology
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Facts about milk

• Milk is a physiological fluid
• Milk is a complete meal
• 4500 different mammals
• Water, protein, lipid, carbohydrate, calcium,
  salts
• Differences among species
• Production of bovine milk 5x108 tons (1.5x107
  tons of protein)
• Denmark 4.4x106 tons of milk

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Lactose
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Convergent Evolution in Lactose Tolerance in
Humans 6000-7000 Years Ago

• Synthesis of lactase in Europeans during life is
  due to mutation in the promoter in the lactase
  gene at position 13910(C/T)
• In African milk-drinking populations mutation at
  positions 14010(G/C), 13915(T/G)and 13907(C/G)

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Mammary Alveolus
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Terminal End Bud
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Milk secretion

• Secretory Pathways
• I Exocytosis - most proteins
• II Fat secretion
• III Transport across PM
• IV Transcytosis
• V Paracellular transport

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Milk Constituents
Serum
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Proteins in bovine milk

• Caseins 80 of total
  protein
• Beta-lactoglobulin 9.8
• Alfa-lactalbumin 3.7
• Serum albumin 1.2
• Immunoglobulins 2.1
• Rest 3-4

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Casein micelles
Carrol et al., 1985
Schmidt, 1982
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McMahon and McManus, 1998
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Formation of cheeseCoagulation of casein with
chymosin
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Milk Constituents
Serum
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Formation of Fat Globules
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Fat Globule Membrane Proteins
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MUC 1

• 580 amino acids
• 11 partially conserved VNTR repeats
• Heavily O-glycosylated

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Mucins
Intestinal epithelium
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Fat Globule Membrane Proteins
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Lactadherin
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Membrane binding C2 domain
Human factor VIII
Lactadherin
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Asymmetry
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ApoptosisProgrammed cell death
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Final stage of apoptosis
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Apoptosis
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Death of rotavirus infection
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Inhibition of rotavirus

• Human MFGM and lactadherin inhibit infection
• Components in bovine milk also inhibit rotavirus
  effectively
• MFGM
• MUC1 (Sialic acid)
• Bovine lactadherin has no effect

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NPC2

• 130 amino acids, three disulphides, mannose 6
  phosphate
• Very resistent to proteolysis
• Approx 20 mg/l milk
• Binds cholesterol
• Essential for intracellular cholesterol
  homeostasis

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Science December 2000
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Filipin-cholesterol staining of cultured human
skin fibroblasts
Norman fibroblasts Fibroblasts from NPC2
patient Fibroblasts from NPC2 patient
NPC2 protein from milk
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Osteopontin

• 262 amino acids, 28 phosphorylations, 3
  O-glycosylations
• RGD-integrin binding sequence
• Phosphorylations are arranged in clusters
• Present in most tissues and body fluids bone,
  plasma, urine, milk
• 20-30 mg/l milk

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Osteopontin functions

• Moderator/regulator of immune functions by
• inducing pro-/antinflammatory cytokine
  production in macrophages and T-cells
  (dose-dependent)
• Calcium transporter binds 50 mol Ca2
• Mediates bone cell binding to hydroxyapatite
• Up-regulated expression in bone growth zones and
  fracture sites.
• Up-regulated in wound healing processes
• Exact function in milk unclear

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a-lactalbumin

• Globular protein
• 123 amino acids
• 8 cysteines
• Homologous with lysozyme
• Cofactor for lactose synthase
• Binds calcium
• Fragments polymerize to stable structures
• Model for molten globule structures

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HAMLET (human alfa-lactalbumin made lethal to
tumor cells)Svanborg et al.

• A complex between apo alfa-lactalbumin and oleic
  acid
• Kills a wide variety of tumor cells but leaves
  normal cells intact
• Treatment of warts
• Effect on bladder cancer
• Effect in a rat Glia blastoma model

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BAMLET (bovine alfa-lactalbumin made lethal to
tumor cells)