Proteins and enzymes are now generally thought to be organized within the c
ell to form clusters in a dynamic and versatile way, and heterologous prote
in-protein interactions are believed to be involved in virtually all cellul
ar events. Therefore we need appropriate tools to detect and study such int
eractions. Chromatographic techniques prove to be well suited for this kind
of investigation. Real complexes formed between proteins can be studied by
classic gel filtration. When enzymes are studied, active enzyme gel chroma
tography is a useful alternative. A variant of classic gel filtration is ge
l filtration equilibrium analysis, which is similar to equilibrium dialysis
. When the association formed is only dynamic and equilibrates very rapidly
, either the Hummel-Dryer method of equilibrium gel filtration or large-zon
e equilibrium filtration sometimes allows the interactions to be analyzed,
both qualitatively and quantitatively. Very often, however, interactions be
tween enzymes and proteins can only be evidenced in vitro in media that mim
ic the intracellular situation. Immobilized proteins are excellent tools fo
r this type of research. Several examples are indeed known where the immobi
lization of an enzyme on a solid support does not affect its real propertie
s, but rather changes its environment in such a way that the diffusion beco
mes limiting. Affinity chromatography using immobilized proteins allows the
analysis of heterologous protein-protein interactions, both qualitatively
and quantitatively. A useful alternative appears to be affinity electrophor
esis. The latter technique, however, is exclusively qualitative. All these
techniques are described and illustrated with examples taken from the liter
ature. (C) 1999 Academic Press.