Post-translational modifications of the core-specific lectin. Relationship to assembly, ligand binding, and secretion.

K.J Colley,J.U Baenziger

doi:10.1016/s0021-9258(18)61112-3

Abstract

The rat core-specific lectin (CSL) or mannan-binding protein is synthesized and secreted by rat hepatocytes and H-4-II-E hepatoma cells. Prior to secretion proline and lysine residues with collagen-like sequences undergo hydroxylation and subsequent glycosylation of hydroxylysine to produce glucosylgalactosylhydroxylysine. Hydroxylation and subsequent glycosylation are inhibited by alpha,alpha'-dipyridyl (Colley, K. J., and Baenziger, U. U. (1987) J. Biol. Chem. 262, 10290-10295). We have used alpha,alpha'-dipyridyl to investigate the role of hydroxylation and glycosylation on interchain disulfide bond formation, assembly of subunits into high molecular weight complexes, attainment of carbohydrate and lipid binding ability, and secretion. Formation of disulfide-bonded dimers and trimers in the endoplasmic reticulum, assembly into high molecular weight complexes in the Golgi, and attainment of carbohydrate binding activity occur in either the presence or absence of these post-translational modifications. The mature fully processed form of the CSL binds hydrophobic matrices and is secreted at a slow, but linear, rate. Inhibition of proline and lysine hydroxylation and hydroxylysine glycosylation prevents CSL secretion and attainment of binding activity for hydrophobic matrices. Secretion of the lectin, although slow, appears to be an active process and may be related to the capacity to interact with membranes and/or lipids. Other proteins known to contain collagen-like sequences such as acetylcholinesterase, pulmonary surfactant apoproteins, and C1q also interact with lipids and/or membranes. The collagen-like domains of these proteins may also play a role in promoting such interactions.

Highlights

The ractore-specific lectin (CSLo)r mannan-binding its ability torecognize features of the “core” region of asparprotein is synthesized and secretedby rat hepatocytes agine-linked oligosaccharides and to bind to yeast mannans, and H-4-11-E hepatoma cells
The core-specific lectin (CSL) is probably identical to the rat and lysinreesiduews itchollagen-likseequences mannan-binding protein (MBP-C) recently characterizedby undergo hydroxylation and subsequent glycosylation Drickamer et al [2]
The mature fully processed form of the CSL binds not account for the prolonged retention of the lectin within hydrophobic matrices and is secreted at a slow, but the cell prior t o secretion [4]

Summary

RESULTS

7.8,200 mM NaCI, 2 mM EDTA, 2% Triton X-loo), andlysates were brought to a finalconcentration of 25mM CaCI,. The dimeric form a,a’-dipyridyl (Fig. 2).CSL was immunoprecipitated from inis detected even in the absence of any chase period and is dividual gradient fractions, and the immunoprecipitatews ere increased inquantity a t 40 min as well. The effect of proline andlysine hydroxylationon interchain disulfide bond formationwas examined inan analogous fashion (Fig. 1).Lysine and proline hydroxylation was prevented by exposing H-4-II-E cells to 3.5 mM a,a’-dipyridyland labeling with [%]methionine for 10 min followed by 0, 10, or 40 min of chase with unlabeled methionine. Hydroxylation of proline and/or lysine is, not essential for theformation of disulfide-bondeddimeric and trimeric formsof the CSL

The formationof interchain disulfide bonds within the first

DISCUSSION

Findings

The properties of pulmonary surfactant proteins and those