Clinical Biochemistry of Domestic Animals (Sixth Edition) - UMK ...

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Chapter | 5 Proteins, Proteomics, and the Dysproteinemias
make a major contribution to the diagnosis of disease.
However, it is a salutary lesson that currently only a limited
number of the many plasma proteins are used for diagnostic
analysis ( Anderson and Anderson, 2002 ).
This chapter focuses on the biochemistry, the diagnostic
methodology, and use in disease diagnosis of measuring
the concentration of serum or plasma proteins, but it
will exclude a number of groups of proteins where the
interpretation of results is more appropriate for other chapters.
Immunoglobulins and complement will be covered
in Chapter 6, lipoproteins in Chapter 4, and fibrinogen in
Chapter 10 . Studies on the proteins of blood have been performed
on serum or plasma. Where appropriate, a distinction
is made between these fluids, although apart from the
absence of fibrinogen in serum, most diagnostic investigations
can be applied to either. Nevertheless serum is the
preferred sample for most of the diagnostic assays used to
investigate the proteins of blood.
II . CLASSIFICATION OF PROTEINS
A . Structural Classification
The structure of proteins is defined by increasing levels
of complexity. The primary structure of a protein is the
sequence of amino acids that makes up the unique composition
of the individual protein. The amino acids are joined
together by peptide bonds linking the carboxylic acid
group of one amino acid to the amino group of the neighboring
amino acid in the chain. With 20 different amino
acids occurring in proteins and with the possibility of more
than a hundred or more amino acids making up the primary
structure of each protein, there are an almost infinite number
of potential proteins that could be present in cells and
tissues. However, the sequence of the amino acids in a particular
protein is predetermined by the order of nucleotide
bases in nuclear DNA, which contains the genetic code for
that protein.
Secondary structure is the presence in protein of regular
structures formed by the linked amino acids giving identifiably
similar three-dimensional conformations. These may
be repeated at intervals in the three-dimensional molecular
structure of the protein. The most important of these structures
are the α -helix and the β -sheet. The α -helix is a righthanded
helix stabilized by hydrogen bonds between the
C O group of one amino acid residue and the N-H group
of another amino acid located four residues along the
peptide chain. The β -sheet is also stabilized by hydrogen
bonds between carboxyl and amino groups of amino acid
residues, but the interacting residues are at different parts
of the same chain. An example of the α -helix is shown by
the structure of albumin (Section IV.A), whereas the contribution
of β -sheets to protein structure is illustrated by
the structure of C-reactive protein (CRP) (Section IV.B.1).
The α -helices and β -sheets can associate together into
supersecondary structures forming recognized motifs
among which the β -meander, Greek key, and β -barrel
structures have been described ( Walsh, 2002 )
The tertiary structure of proteins is the three-dimensional
structure of the protein and is dependent on its primary and
secondary structures. This native conformation of the protein
is essential for its activity and depends on the correct
folding of the protein after synthesis. Most proteins above
a certain size can be subdivided into domains, which are
independent folding units. The conformation of the protein
is held together by weak forces between amino acid
side chains such as hydrogen bonds, electrostatic and
hydrophobic interaction, and also by covalent disulphide
bonds between cysteine residues. Great strides have been
made in determining the structure of proteins using X-ray
crystallography and nuclear magnetic resonance (NMR).
Structures of many proteins, including serum protein, can
be obtained from online, open-access databases such as
the Research Collaboratory for Structural Bioinformatics
Protein Data Base located on the Internet at www.rcsb.org/
pdb. The structures can be manipulated by protein modeling
software, among which Protein Explorer or RasMol can
be downloaded from www.umass.edu/microbio/rasmol.
The quaternary structure of proteins is the combination
of protein subunits to create a multisubunit complex.
Thus, hemoglobin requires the combination of four subunits
(two α -chains and two β -chains) into a tetramer for
the fully functional protein. Examples of serum proteins
that have quaternary structure include immunoglobulins,
formed from two light chains and two heavy chains, and
CRP, in which five subunits combine to form a pentameric
structure.
A further classification of protein based on their structure
is between “ fibrous ” and “ globular ” proteins. The
former adopt elongated three-dimensional shapes in their
quaternary structure and are usually involved in structural
roles in biological systems such as α -keratin, collagen, and
elastin. Apart from fibrinogen (Section VI.B.5), which has
as its function the formation of fibrin, fibrous proteins are
not found in plasma protein. Thus, the majority of plasma
proteins are globular proteins , adopting complex threedimensional
shapes by folding of the polypeptide chain.
B . Chemical Classification
Proteins are also classified as “simple ” or “conjugated. ”
Simple proteins contain only a polypeptide chain of linked
amino acids, whereas conjugated proteins contain nonamino
acid components. These can be carbohydrate residues (glycoprotein
and proteoglycan), metal ions such as Fe 2 or
Ca 2 (metalloproteins), phosphate (phosphoproteins), lipid
(lipoproteins), and nucleic acids (nucleoproteins such as
histones) . Many plasma proteins are conjugated to carbohydrate
and are present in the circulation as glycoproteins.