Food Chemistry

1.4 Proteins 41

which determines the molecular conformation
(secondary and tertiary structures). Proteins
sometimes occur as molecular aggregates which
are arranged in an orderly geometric fashion
(quaternary structure). The sequences and con-
formations of a large number of proteins have
been elucidated and recorded in several data
bases.

(1.82)

Glycoproteins, such asκ-casein (cf. 10.1.2.1.1),
various components of egg white (cf. 11.2.3.1)
and egg yolk (cf. 11.2.4.1.2), collagen from
connective tissue (cf. 12.3.2.3.1) and serum
proteins of some species of fish (cf. 13.1.4.2.4),
contain one or more monosaccharide or oligosac-
charide units bound O-glycosidically to serine,
threonine orδ-hydroxylysine or N-glycosidically
to asparagine (Formula 1.82). In glycoproteins,
the primary structure of the protein is defined
genetically. The carbohydrate components,
however, are enzymatically coupled to the
protein in a co- or post-transcriptional step.
Therefore, the carbohydrate composition of
glycoproteins is inhomogeneous (microhetero-
geneity).

1.4.1 AminoAcidSequence

1.4.1.1 AminoAcidComposition,Subunits........................

Sequence analysis can only be conducted on

a pure protein. First, the amino acid compo-

sition is determined after acidic hydrolysis.

The procedure (separation on a single cation-

exchange resin column and color development

with ninhydrin reagent or fluorescamine) has

been standardized and automated (amino acid

analyzers). Figure 1.10 shows a typical amino

acid chromatogram.

As an alternative to these established methods,

the derivatization of amino acids with the sub-

sequent separation and detection of derivatives

is possible (pre-column derivatization). Various

derivatization reagents can be selected, such as:

• 9-Fluorenylmethylchloroformate
  (FMOC, cf. 1.2.4.2.1)


• Phenylisothiocyanate (PITC, cf. 1.2.4.2.3)


• Dimethylaminoazobenzenesulfonylchloride
  (DABS-Cl, cf. 1.2.4.2.1)


• Dimethylaminonaphthalenesulfonylchloride
  (DANS-Cl, cf. 1.2.4.2.1)


• 7-Fluoro-4-nitrobenzo-2-oxa-1,3-diazole
  (NBDF, cf. 1.2.4.2.1)


• 7-Chloro-4-nitrobenzo-2-oxa-1,3-diazole
  (NBDCl, cf. 1.2.4.2.1)


• o-Phthaldialdehyde (OPA, cf. 1.2.4.2.4)

It is also necessary to know the molecular weight

of the protein. This is determined by gel column

chromatography, ultracentrifugation or SDS-

PAG electrophoresis. Furthermore, it is necessary

to determine whether the protein is a single

molecule or consists of a number of identical or

different polypeptide chains (subunits) associated

through disulfide bonds or noncovalent forces.

Dissociation into subunits can be accomplished

by a change in pH, by chemical modification of

the protein, such as by succinylation, or with de-

naturing agents (urea, guanidine hydrochloride,

sodium dodecyl sulfate). Disulfide bonds, which

are also found in proteins which consist of only

one peptide chain, can be cleaved by oxidation of

cystine to cysteic acid or by reduction to cysteine

with subsequent alkylation of the thiol group