Abstract
Histidine-Proline-rich Glycoprotein (HPRG) is a plasma protein of vertebrates and several marine bivalves. Due to its multidomain structure consisting of several regions HPRG can interact with a variety of ligands, however the exact physiological role has not been discovered yet. Past purification approaches out of plasma or serum often led to co-purification of other proteins so that for a profound understanding of the function it is important to obtain a protein of high purity. Recent purification strategies were based upon metale chelate affinity chromatography followed by anion exchange chromatography or size exclusion chromatography, respectively. A large amount of serum albumin, the major plasma protein, also elutes from metale chelate affinity chromatography columns. Separation of rabbit HPRG from rabbit serum albumin could not be achieved via the above named methods by us. We present a method of purification of rabbit serum HPRG by means of metal affinity chromatography and preparative gel electrophoresis, which makes it possible to obtain HPRG practically devoid of impurities as assessed by mass spectrometry analysis. Moreover, we characterize the amount of glycosylation of HPRG and–to the best of our knowledge for the first time–the glycosylation pattern of rabbit HPRG.
Highlights
Histidine-Proline-rich Glycoprotein (HPRG), or alternatively Histidine-rich Glycoprotein (HRG), is a protein of vertebrate plasma [1, 2] that was recently found in the plasma of several marine bivalves [3, 4]
HPRG is a plasma glycoprotein that occurs in many vertebrates
In our case we were not able to separate the rabbit serum albumin, which did elute during metal chelate affinity chromatography
Summary
Histidine-Proline-rich Glycoprotein (HPRG), or alternatively Histidine-rich Glycoprotein (HRG), is a protein of vertebrate plasma [1, 2] that was recently found in the plasma of several marine bivalves [3, 4]. First isolated and characterized in 1972 by Heimburger et al [5, 6] the concentration in human plasma was found to be 100–150 mg/l, whereas in rabbit plasma a concentration of up to 900 mg/l was demonstrated [7]. HPRG, alongside fetuins and kininogen, is a type 3 cystatin and a member of the cystatin superfamily. Like most of the other members of the type 3 subgroup HPRG is produced in the liver [8,9,10,11]
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