Aminoalkyl matrices are used in affinity chromatography of amine oxidases a
nd other proteins with affinity for amino groups. Under appropriate circums
tances chromatography on aminoalkyl matrices may yield purification factors
around 100 to 1000, and they have been used in affinity purification of ma
ny members of the amine oxidase family.
Other proteins with affinity for aminoalkyl matrices include thiol ester pr
oteins, lactoferrin, and proteins with lysine-binding kringles (plasminogen
, plasminogen activator, apolipoprotein A).
The affinity of thiol ester proteins for aminoalkyl matrices is abolished a
fter inactivation of the thiol ester group by reaction with low molecular w
eight amines including ammonia. Due to this, an ammonium sulphate precipita
tion step should be included in purification schemes for amine oxidases.
The affinity of lactoferrin for aminoalkyl matrices stems from an affinity
for the repeating amino groups in glycosaminoglycans, and this explains why
lactoferrin requires diamines for efficient elution.
The affinity of plasminogen for aminoalkyl groups is exploited in a one-ste
p purification from plasma, and is also utilised in purification schemes fo
r angiostatin, an angiogenesis-inhibiting fragment of plasminogen. Apolipop
rotein A is homologous to plasminogen, and also has affinity for aminohexyl
columns. The common binding motif for these proteins are lysine-binding kr
ingles.
Due to the properties of the amino group itself, aminoalkyl matrices will i
nevitably also function as anion exchangers, and this must be taken into co
nsideration in the choice of conditions for sample loading, column washing
and elution of bound proteins. Depending on the length of the alkyl chain,
the matrices also have a potential for hydrophobic interactions. This prope
rty has been exploited in the purification of several proteins but must be
minimized during affinity chromatography of amine oxidases.
In conclusion, aminoalkyl matrices are valuable tools for affinity chromato
graphy of several different proteins, and simple variations of sample pretr
eatment, sample loading, and column washing and elution conditions allow ef
ficient selective purification of proteins with different affinities for th
e matrices. (C) 2001 Elsevier Science B.V. All rights reserved.