Interaction of bovine gallbladder mucin and calcium-binding protein: effects on calcium phosphate precipitation.
OBJECTIVE
Gallstones consist of calcium salts and cholesterol crystals, arrayed on a matrix of gallbladder mucin (GBM), and regulatory proteins like calcium-binding protein (CBP). To determine if interactions between CBP and GBM follow a biomineralization scheme, their mutual binding and effects on CaHPO4 precipitation were studied.


METHODS
Binding of CBP to GBM was assessed by inhibition of the fluorescence of the complex of GBM with bis-1,8-anilinonaphthalene sulfonic acid (bis-ANS). The effects of the proteins on precipitation of CaHPO4 were assessed by nephelometry and gravimetry. Precipitates were analyzed for calcium, phosphate, and protein.


RESULTS
CBP and bis-ANS competitively displaced each other from 30 binding sites on mucin, with a 1:1 stoichiometry and similar affinity. The rate of precipitation of CaHPO4 was retarded by mucin and CBP. Precipitate mass was unaffected by GBM alone but decreased with the addition of CBP. Complexing CBP with GBM abolished or moderated this latter effect, altered precipitate morphology, and changed the stoichiometric ratios of Ca to PO4 in the precipitates from 1:1 to 3:2. Mucin and CBP were incorporated into the precipitates.


CONCLUSIONS
These studies suggest that the formation of calcium-containing gallstones is a biomineralization process regulated by both GBM and CBP.
Afdhal
N H
NH
Section of Gastroenterology, Thorndike Memorial Laboratories, Evans Department of Medicine, Boston City Hospital, Massachusetts, USA.
Ostrow
J D
JD
Koehler
R
R
Niu
N
N
Groen
A K
AK
Veis
A
A
Nunes
D P
DP
Offner
G D
GD
D000817:Anilino Naphthalenesulfonates; D000818:Animals; D001667:Binding, Competitive; D002130:Calcium Phosphates / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000502:physiology*; D002417:Cattle; D011232:Chemical Precipitation; D002769:Cholelithiasis / Q000378:metabolism; D005456:Fluorescent Dyes; D005704:Gallbladder / Q000378:metabolism*; D066298:In Vitro Techniques; D009077:Mucins / Q000502:physiology*; D009391:Nephelometry and Turbidimetry
D000817:Anilino Naphthalenesulfonates; D002130:Calcium Phosphates; D002135:Calcium-Binding Proteins; D005456:Fluorescent Dyes; D009077:Mucins; C485828:alpha-tricalcium phosphate; C485830:tetracalcium phosphate; C035763:5,5'-bis(8-(phenylamino)-1-naphthalenesulfonate); C485838:calcium phosphate, monobasic, anhydrous; C020243:calcium phosphate; C485829:calcium phosphate, dibasic, anhydrous