The pronase resistance of cholesterol-nucleating glycoproteins in human bile.
OBJECTIVE
Many putative pronucleating proteins have been isolated from the biliary concanavalin A (con A)-binding fraction. The pronase resistance of the overall nucleating-promoting activity was almost never taken into consideration. The aim of this study was to identify the major pronase-resistant con A-binding glycoproteins.


METHODS
Pronase-treated and -untreated con A-binding glycoproteins were separated on a Superose 12 gel permeation column (Pharmacia, Uppsala, Sweden) and tested in a crystal growth assay. Proteins were identified by amino-terminal sequencing.


RESULTS
Con A-binding pronucleating activity eluted in two peaks on the Superose column. This activity was unaltered after pronase treatment. Activity peak I contained too little protein to allow amino-terminal sequencing. In activity peak II, the major pronase-resistant con A-binding glycoproteins were identified as alpha 1-antitrypsin and alpha 1-antichymotrypsin. The 130-kilodalton nucleation promoter was identified as aminopeptidase N, but the full pronase resistance of this protein, reported earlier, was not confirmed. Immunoabsorptive removal of alpha 1-antitrypsin and alpha 1-antichymotrypsin and immunopurification showed that only alpha 1-antichymotrypsin had pronucleating activity.


CONCLUSIONS
The pronase resistance of the nucleating-promoting activity of the con A-binding glycoprotein fraction was confirmed. An important part of this activity could be attributed to alpha 1-antichymotrypsin. It is an acute-phase protein, as are many other pronucleating proteins, which might indicate a general mechanism of action in gallstone formation.
Zijlstra
A I
AI
Department of Gastroenterology and Hepatology, Academic Medical Center, Amsterdam, The Netherlands.
Offner
G D
GD
Afdhal
N H
NH
van Overveld
M
M
Tytgat
G N
GN
Groen
A K
AK
D000328:Adult; D000368:Aged; D000595:Amino Acid Sequence; D001646:Bile / Q000187:drug effects / Q000378:metabolism*; D002784:Cholesterol / Q000502:physiology*; D002850:Chromatography, Gel; D003208:Concanavalin A; D003460:Crystallization; D004351:Drug Resistance; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006023:Glycoproteins / Q000235:genetics / Q000502:physiology*; D006801:Humans; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D011402:Pronase / Q000235:genetics / Q000494:pharmacology*; D012685:Sepharose; D000514:alpha 1-Antichymotrypsin / Q000502:physiology; D000515:alpha 1-Antitrypsin / Q000502:physiology
D006023:Glycoproteins; D000514:alpha 1-Antichymotrypsin; D000515:alpha 1-Antitrypsin; D003208:Concanavalin A; D012685:Sepharose; C049225:Superose 12; D002784:Cholesterol; D011402:Pronase