Amino Acid Metabolism in the Mammary Gland

Proteins are composed of amino acids. Amino acid utilization by mammary tissue is affected by
a) blood concentrations of amino acid,
b) mechanisms of amino acid uptake into mammary cells, and
c) intracellular metabolism of the amino acid.
Each of these levels of control is further influenced by a range of factors. Blood amino acid
concentrations are affected by nutrition, but also by physiological state of the animal and other
factors.

In terms of intracellular metabolism of amino acids, once inside the cell, AA's may:

1. undergo mRNA-directed polymerization to form milk proteins.

2. enter into metabolic reactions yielding carbon dioxide, urea, polyamines, and
nonessential AA's (NEAA's)
3. be retained in cells as part of structural proteins and enzymes
4. pass unchanged into milk, blood or lymph

Routes 3 and 4 are minor in the mammary gland relative to the tremendous synthesis and
secretion of milk proteins.

Proteins constitute 50% or more of the dry weight of a cell. They include:

 Enzymes - There are a large number of enzymes involved in milk synthesis. For example,
lactose synthase is responsible for lactose synthesis, acetyl-CoA carboxylase and fatty
acid synthetase are two type of enzymes involved in milk aft synthesis, and several
proteases are involved in milk protein processing and interactions of the mammary cels
with their extraccellular environment.
 Structural proteins - There are many structural proteins involved in mammary gland
structure and function. For example, collagens, laminin and other connective tissue
proteins form the basis of the extracellular matrix of the tissue (the noncellular
components of the stroma and the basement membranes of the alveoli), and cytoskeletal
proteins are involved in cell structure, cell movement and cell function (tubulin, actin,
cytokeratin are some discussed in the Histology and Cell Biology section of the
Mammary Structure Lesson).
 Nutrients - Casein is a good example of a protein which supplies nutrients (amino acids).
 Hormones - Prolactin, growth hormone, placental lactogen, insulin, and a number of
growth factors are examples of hormones and growth factors which affect mammary
gland function. Oxytocin is a peptide hormone which causes myoepithelial cell
contractions during milk ejection.
 Protective agents - Immunoglobulins, lactoferrin (an iron-binding protein), lysozyme (an
enzyme), and lactoperoxidase (another enzyme) are a few examples of factors which can
protect the mammary gland from infection.
 Toxins - Some bacterial toxins are peptides or derivatives of peptides. These may cause
inflammation in the mammary tissue.
  Others - Transport of substrates into the mammary cell occurs by protein molecules
associated with the cell plasma membrane; transcription factors in the nucleus and
translation factors in the cytoplasm are proteins; hormone receptors and growth factor
receptors are proteins; and there are many more.

Major Milk Proteins

All of the major milk proteins (except serum albumin and immunoglobulins) are synthesized by
epithelial cells in the mammary gland from amino acids (AA's) extracted from the blood.
Approx.
% of Isoelectric Molecular 
Protein skim
milk point weight
protein
alpha-Casein 45-55 4.1 23,000
kappa-Casein 8-15 4.1 19,000
beta-Casein 25-35 4.5 24,000
gamma-Casein 3-7 5.8-6.0 --
alpha-
2-5 5.1 14,437
Lactalbumin
beta-
7-12 5.3 18,000
Lactoglobulin
Blood serum
0.7-1.3 4.7 68,000
albumin
Lactoferrin 0.2-0.8  -- 87,000
Immunoglobulins
  --  --  --
:
IgG1 1-2  -- 160,000
IgG2 0.2-0.5  -- 160,000
~1,000,00
IgM 0.1-0.2  --
0
IgA 0.05-0.10  -- ~400,000
Proteose 4,100 to
2-6 3.3-3.7
peptone fraction 200,000

Caseins (CN):
Found in milk in the form of a Micelle (a dense protein granule). The phosphate groups
covalently bound to the casein molecules are involved in binding Calcium (Ca) via ionic bonds.
After caseins are phosphorylated, Ca++ binds to the phosphate to initiate polymerization of the
micelle particles. This [casein-PO4- Ca++ - PO4-casein] structure is key to micelle formation.
The casein micelle functions as a source of nutrients for the neonate: supplying amino acids,
calcium and phosphate.

Micelles are ~140 nanometers in diameter. They are composed of alpha- (as-), beta-, and kappa-
caseins. a-Caseins are in multiphosphorylated forms (as2, as3, as4, as5, and as6). ß-Casein is a
major casein in cow milk, but is the minor casein in human milk. k-Casein (a glycoprotein) is
distributed throughout the casein micelle and acts to stabilize the micelle. Gamma-caseins (part
of the proteose peptone fraction of milk) are C-terminal fragments of ß-casein, which are
released by plasmin digestion, mostly while the milk is in the gland.

The destabilization of the casein micelle structure and partial hydrolysis of casein decreases the
quality of fluid milk and the yield of cheese from milk. Conversely, the destabilization of the
casein micelle by proteases is part of the mechanism involved in milk digestion in the stomach
and intestine. Controlled hydrolysis of casein is also the means of producing cheeses and other
cultured milk products.

Major Whey Proteins:

ß-LACTOGLOBULIN (ß-LG): Is ~50 % of the total whey protein content in milk. Is the major
whey protein in ruminants and pigs. It is not found in milk of many species. The function of ß-
LG is unknown; it may be a fatty acid or lipid binding protein. It does have sequence similarities
with retinol-binding proteins, but this may not be its function. Generally it is found in milk of
species which transport high levels of immunoglobulins during colostrum formation, however
the specific relationship between the presence of ß-LG and immunoglobulin transport remains
unclear.

a-LACTALBUMIN (a-LA): Is ~25 % of the total whey protein content in milk. Is the "B"
protein of the Lactose Synthase enzyme complex (see Lactose Synthesis Lesson). Therefore, it is
of major interest in terms of the control of milk secretion. May have other nonspecific effects on
the integrity of milk fat membranes. a-LA binds Ca and Zn.

Minor Whey Proteins:

SERUM ALBUMIN (SA) : Serum albumin comes from the serum; it is not synthesized in the
mammary gland. It is presumed to enter the milk via "leakage" by the paracellular pathway, or
by uptake with other components such as immunoglobulins. There does not seem to be a more
specific mechanism of transport. Increases in milk concentration of serum albumin occur
especially during mastitis and during mammary involution. Function of serum albumin in milk is
unknown. It does bind to fatty acids, as well as other small molecules.

IMMUNOGLOBULINS (Ig): include IgG1, IgG2, IgA, IgM : Immunoglobulins are in very high
concentrations in colostrum, but much lower concentrations in milk. Immunoglobulins are part
of the passive immunity transported to the neonate via colostrum in many species. They are part
of the mammary immune system. Secretory Component (SC) is the part of the IgA receptor
which hydrolysed by a protease and remains attached to IgA as it is secreted from the cell.

Other Milk Proteins

Lactoferrin (LF) - Is an iron binding protein and has antibacterial properties. Relatively low
concentrations during lactation in cow milk, but is increased during mastitis and involution.
Lactoferrin may also be an immunomodulator. Lactoferrin is in human milk in high
concentrations; in fact it is the major whey protein in human milk. Lactoferrin is the major
nonspecific disease resistance factor found in the mammary gland. [See also the sections on Dry
Period and Mammary Involution and Mastitis).

Lactoperoxidase - Is an enzyme that breaks down hydrogen peroxide. It has antibacterial

properties.

Lysozyme - Is an enzyme that cleaves the carbohydrate polymers of the bacterial cell wall. High
activity in human milk and possibly in horse milk, but very low activity in cow milk.

ß2-Microglobulin - Initially it was discovered as a crystalline precipitate in resuspended casein

and initially was called lactolin. It is now know to be part of the Major Histocompatability
Complex II (MHC II). While the function in milk is not known, it more recently has been found
in the epithelial cell to be associated with a protien that binds to immunoglobulin G. It is thought
that ß2-Microglobulin may be involved in the functionality of that IgG receptor in the process of
IgG transport during colostrum formation. This latter function for ß2-Microglobulin remains
controversial.

Milk has numerous other enzymes including: proteases, protease activators, nucleases,
glycosidases, and others

Skim milk has cell membranes and their associated membrane-bound enzymes such as
galactosyltransferase (the "A" protein component of Lactose Synthase; see the Lactose Synthesis
section ).

Milk Proteins and Their Synthesis

The proteins that will be secreted by the cell, including the milk proteins synthesized by the
epithelial cell, are synthesized in the rough endoplasmic reticulum. Kappa-casein also is initially
glycosylated in the RER. Proteins are then transported to the Golgi apparatus where they are
further processed. For example, the caseins are phosphorylated and the micelles are formed in
the Golgi. Remember from the sections on lactose that it also is synthesized in the Golgi
apparatus.