Influence of thyroxine in the regulation of rat parotid salivary protein composition

This study tested the role of thyroxine in the regulation of the protein composition of rat parotid saliva. Thyro-parathyroidectomy was performed on two groups of rats, one of which subsequently received thyroxine replacement; the third group was sham-operated. Parotid saliva was collected on the eighth day after surgery, with pilocarpine and isoproterenol used as a secretory stimulus. The volume of saliva collected in 30 min from the thyro-parathyroidectomized rats was 55% less than that collected from sham-operated rats. In the thyro-parathyroidectomized rats, the protein concentration as measured by absorption at 215 nm was unaltered, but that measured by the Lowry procedure was 43% higher. Spectrophotometric scans of Coomassie Blue-stained gels following sodium dodecylsulfate polyacrylamide gel electrophoresis of the secreted proteins showed an 18% reduction in the proportion of protein attributable to amylase and a 43% reduction in proportion of acidic and basic proline-rich proteins following thyro-parathyroidectomy; deoxyribonuclease and two other major secretory proteins (Fraction I and Fraction V) were increased (38%, 20%, and 46%, respectively). These changes in flow rate, protein concentration by the Lowry assay, and protein composition were prevented by treatment of thyro-parathyroidectomized rats with thyroxine replacement and are in opposition to those changes we reported earlier for hyperthyroid rats. The results indicate that the flow of saliva as well as the synthesis of the various salivary proteins are influenced by thyroxine.     

Changes in rat parotid saliva protein composition following chronic reserpine treatment and their relation to inanition

Chronic administration of the catecholamine-depleting agent, reserpine (0.5 mg/kg), resulted in a reduction in food intake after 3 days. To differentiate effects of the drug from those of reduced food intake a pair-fed group, whose daily caloric intake was restricted to the amount consumed by the reserpine-treated rats, was included. After 7 days, both the reserpine-treated and pair-fed control exhibited a marked reduction in the volume of saliva collected in a 30 min interval following a secretory stimulus compared to untreated ad libitum-fed controls, and the proportion of salivary proteins attributable to acidic and basic proline-rich proteins and to minor 1b protein were decreased whereas deoxyribonuclease was increased. For two of the salivary proteins (fractions I and V) changes for the reserpine-treated and pair-fed groups were different. Fraction I was reduced in both groups, but exhibited a greater decrease in the pair-fed than in the reserpine-treated, whereas fraction V was significantly increased only in the pair-fed group. Thus many of the salivary changes associated with reserpine treatment may have resulted from the change in feeding habits and not from reserpine treatment per se. The study demonstrates the importance of controlling for food intake under experimental circumstances which may lead to a marked change in daily feeding habits.     

Regulation of salivary proteins

Previous studies have shown that several factors--such as alloxan-induced diabetes, adrenalectomy, or removal of the thyroid-parathyroid gland complex--can influence the flow rate, protein concentration, and protein composition of rat parotid saliva. The present study was undertaken to explore further the influence of glucocorticoids and thyroxine on rat parotid saliva in hormonally intact animals. As compared with untreated animals, adult male rats treated with 10 micrograms dexamethasone per 100 g body weight for eight days demonstrated a 75% reduction in volume of parotid saliva secreted in response to a uniform stimulus. The protein concentration of the saliva was increased three-fold. Sodium dodecyl sulfate polyacrylamide gel electrophoresis showed relative decreases in acidic and basic proline-rich proteins and in a protein identified as Fraction V, while amylase was increased. The electron microscopic appearance of the granules was markedly different from that of the control, in that the granules exhibited an electron-dense periphery and core, with the remainder of the granule having an electronlucent appearance. In contrast, rats treated for eight days with 20 micrograms thyroxine per 100 g body weight exhibited a 50% increase in volume of saliva collected in response to a secretory stimulus. Although the concentration of protein was not different from that of the control, gel electrophoresis showed relative increases in acidic and basic proline-rich proteins and a decrease in Fraction V. Amylase was unchanged. The secretory granules of thyroxine-treated rats were electronlucent and amorphous. The granules appeared to coalesce within the cell.(ABSTRACT TRUNCATED AT 250 WORDS)     

Changes in rat parotid salivary proteins associated with liquid diet-induced gland atrophy and isoproterenol-induced gland enlargement

The composition of secretory proteins in the parotid saliva of untreated rats, fed a stock pelleted diet (CON), was compared to that of rats maintained on a liquefied diet (LIQ), so as to reduce gland secretory activity, and to rats treated chronically with isoproterenol (ISO), so as to enhance activity. Each experimental treatment resulted in marked changes in protein composition. In CON rats, the basic and acidic proline-rich proteins accounted for 25 per cent of all secretory protein. In LIQ rats, the proportion was 13 per cent. Several basic proline-rich proteins were absent and the major acidic proline-rich protein was markedly reduced. Amylase was reduced whereas DNase and a leucine-rich protein (fraction I) were increased. The proportion of cystine-rich protein (fraction V) was not different from CON rats. The changes observed in the saliva of ISO rats were in marked contrast to these findings. Basic and acidic proline-rich proteins were increased and accounted for 90 per cent of all secretory protein, amylase was responsible for the remaining 10 per cent. Thus the composition of secretory proteins in the parotid saliva of rats can be altered by experimental conditions which affect gland secretory activity. The mechanisms by which these changes occur is not known.     

The effect of chronic atropine treatment on salivary composition and caries in rats

Many instances of salivary dysfunction in humans can be traced to the use of medications that have hyposalivary side-effects. In this study, atropine, a cholinergic antagonist, was administered chronically to rats by use of osmotic mini-pumps. Steady-state blood levels, similar to levels obtained in human multiple oral dosing, were thus maintained. Atropine delivered in this manner for 24 days was found to decrease protein concentration of parotid saliva (p less than 0.05) elicited by pilocarpine, and to increase smooth-surface caries scores (p less than 0.05) in rats fed a cariogenic diet. Parotid saliva collected via ductal cannulation from rats subjected to chronic atropine administration (and stimulated to secrete by pilocarpine) exhibited increased levels of two basic proline-rich proteins (Peak A and SP-3), as evaluated by SDS-PAGE, compared with those observed in saliva from controls. Cannulation of sublingual glands in animals receiving high doses of atropine produced no measurable secretion upon pilocarpine stimulation. Carbachol stimulation of dispersed cell aggregates of sublingual glands from sham-operated and high-dose atropine groups indicated that the glands responded similarly once the antagonist was washed from the system, implying that the lack of secretion in vivo was caused by antagonism of the cholinergic receptor by atropine. Our observations suggest that this model system can be exploited for determination of the effects of chronic administration of hyposalivary drugs on salivary composition and caries rates.     

The effect of chronic propranolol treatment on salivary composition and caries in the rat

Many drugs are known to affect salivary secretion. The purpose of this study was to explore the chronic effects of a commonly used beta-adrenergic blocker, propranolol. Adult rats were desalivated or treated for 28 days with propranolol HCl (10 or 20 mg/kg, daily) or sterile buffer (sham-operated control) using osmotic pumps for delivery. The parotid and submandibular glands of each rat were cannulated and secretion elicited by pilocarpine (10 mg/kg, intravenous). There were no statistical differences in salivary protein content (Lowry) or output among the groups (ANOVA, p greater than 0.05). Analysis of salivary proteins by SDS-PAGE revealed a constant profile for submandibular secretions, but peak A and SP-3 proline-rich proteins were not detectable in parotid saliva of animals treated with propranolol for the entire experiment. Significantly increased smooth-surface (p = 0.0003) and sulcal (p = 0.0011) caries scores were found within these propranolol groups (ANOVA). The findings provide further evidence that chronic administration of propranolol alters salivary composition by decreasing proline-rich proteins and concurrently enhances susceptibility to caries.     

Altered bacterial aggregation and adherence associated with changes in rat parotid-gland salivary proteins induced in vivo by beta-adrenergic stimulation

Reduced adherence and aggregation were associated with protein alterations in parotid saliva after chronic treatment with the beta-adrenergic agonist isoproterenol. In contrast, saliva from animals treated with the beta-antagonist, propranolol, did not cause such changes; the protein composition of this saliva was similar to that of controls. SDS-polyacrylamide gel electrophoresis of protein in saliva samples before and after they were mixed with 10 mg of spheroidal hydroxyapatite beads (HA), as well as protein adsorbed and recovered from the HA, showed that an acidic, proline-rich protein with a molecular weight of approx. 40,000 was the predominant protein adsorbed. This protein was significantly diminished in saliva from isoproterenol-treated rats. Proteins with molecular weights between 44,000 and 48,000 and unique to the saliva from isoproterenol-treated animals were also adsorbed to HA. Thus alterations in proline-rich proteins of parotid saliva may influence the adherence and aggregation of oral bacteria, two processes considered important for in-vivo colonization of oral surfaces.     

Salivary gland function in persons with ectodermal dysplasias

Ectodermal dysplasias (EDs) constitute a group of conditions comprising developmental defects in two or more of the following tissues: hair, teeth, nails, and sweat glands. The aim of the present study was to contribute to a better understanding of salivary gland involvement in EDs. An ED group (n = 39, median age 12 yr; 24 males, 15 females) and a healthy age- and sex-matched control group were studied. Citric acid stimulated submandibular and parotid salivary flow rates and salivary concentrations, and output of total protein, acidic proline-rich proteins and histatins were analysed. The associations between quantitative and qualitative salivary parameters were also studied. In the ED group, 13 persons (33%) demonstrated a significantly reduced secretion of submandibular and/or parotid saliva, in addition to a low unstimulated and/or chewing-stimulated whole salivary flow. In the ED group as a whole, a reduced median secretory rate of submandibular saliva was found, whereas the median concentrations of some protein parameters were increased. However, the overall output of proteins was normal or reduced. Submandibular glands seemed to be more affected than parotid glands in EDs. In conclusion, salivary secretory tests are recommended in persons with known or suspected EDs.     

The association of basic proline-rich peptides from human parotid gland secretions with caries experience

To address whether there are associations between the peptide composition of human parotid saliva and dental decay (caries) experience, we have characterized the peptides from parotid ductal saliva collected from nine adults who have remained free from dental caries (mean age = 59.2; Decayed Missing Filled Surfaces index [DMFS] = 0) and nine individuals who have experienced caries (mean age = 51.2; mean DMFS = 38.4). Ethanol-soluble peptides were size-fractionated on columns of Bio-Gel P-2; the salivary peptides derived from caries-susceptible subjects appeared larger than those found in the saliva of caries-free subjects. Peptides were then resolved into 19 species by cation exchange HPLC. Sequence analysis identified 18 peptides that appear to be proteolytic cleavage products of the basic proline-rich proteins IB-4, IB-5, IB-7, IB-8b, and P-B. The peptides that were more abundant in saliva obtained from the caries-free group differed from those isolated from the caries-susceptible group. The median peptide concentration of one possible precursor protein, IB-7, was found to be higher in saliva collected from caries-free individuals than in that from caries-susceptible individuals. Although differences were found in the phenotypes of proline-rich proteins expressed by these groups of caries-free and caries-susceptible subjects, no statistically significant associations were observed among proline-rich phenotypes and the level of any peptide. Collectively, our results indicate that proteolytic processing of parotid salivary proteins differs among individuals who have remained caries-free and those who have experienced dental decay.     

Isolation and characterization of the basic proline-rich proteins from rat parotid saliva

Five fractions of basic proline-rich proteins were isolated from rat parotid saliva, obtained by surgical cannulation of the ducts. The purification procedures employed DEAE-Sephadex to isolate a heterogeneous break-through fraction containing the basic proline-rich proteins, followed by gel filtration on Sephadex G-200 to separate the high molecular weight glycoprotein, fraction A, from the other basic proline-rich proteins which were resolved into four additional fractions, SP-1 to SP-4, by ion exchange chromatography on SP-Sephadex. The proteins differed in their amino acid composition and content of neutral and amino sugars. All the proteins were characterized by a high proportion of proline (approx. 40 mol per cent) and glycine (11–23 mol per cent). Four of the fractions were also enriched in glutamic $\text{acid}\text{glutamine}$ (19–26 mol per cent). The exception was fraction SP-4, which contained lower levels of glutamic $\text{acid}\text{glutamine}$ and has no counterpart in human basic proline-rich proteins. Fraction A, the basic glycoprotein, was heavily glycosylated (59 mol per cent), whereas SP-2 and SP-4 were less glycosylated. Fractions SP-1 and SP-3 contained low levels of neutral and amino sugars. Basic proline-rich proteins constitute a smaller percentage of the total protein in rat parotid saliva than they do in human parotid saliva (10.5 versus 40 per cent). Rat basic glycoprotein fraction constitutes less than 1 per cent whereas the human glycoprotein fraction constitutes 17 per cent. Rat basic proline-rich proteins appear to be larger and less basic than most of the human basic proteins, and they resolve into fewer protein fractions (4 versus 9) with SP-Sephadex chromatography.     

Salivary non-immunoglobulin agglutinin inhibits human leukocyte elastase digestion of acidic proline-rich salivary proteins

Saliva contains acidic proline-rich salivary proteins that are involved in the formation of the salivary pellicle coating supragingival tooth surfaces. However, human leukocyte elastase, arriving in gingival exudates from inflamed periodontal tissues, degrades the acidic proline-rich salivary proteins, preventing binding to hydroxylapatite surfaces. Here it is reported that high-molecular-weight non-immunoglobulin salivary agglutinin inhibited the proteolytic action of human leukocyte elastase on purified acidic proline-rich salivary proteins. Inhibition was eliminated with monoclonal antibody to a protein determinant on the salivary agglutinin. The addition of antibody against salivary agglutinin blocked the inhibitory effect of parotid saliva on exogenously applied human leukocyte elastase, allowing for the elastase-mediated digestion of the salivary acidic proline-rich salivary proteins. Salivary agglutinin, therefore, is a physiologically important inhibitor of human leukocyte elastase and is able to inhibit elastase-mediated digestion of salivary acidic proline-rich proteins.     

Differences in basic proline-rich proteins in rat parotid saliva following chronic isoproterenol treatment or maintenance on a liquid diet

The basic proline-rich proteins (BPRP) in the stimulated parotid saliva of rats treated for 8 days with isoproterenol and rats fed a liquid diet for 2 weeks were compared to those in the stimulated parotid saliva of untreated rats fed a stock pelleted-diet (control). In the control, the BPRP were separated into 5 groups designated Peak A (the basic proline-rich glycoprotein), SP-1, SP-2, SP-3 and SP-4. The percentage of BPRP in each group was as follows: Peak A, 6.5 per cent; SP-1, 37 per cent; SP-2, 6.5 per cent; SP-3, 32.4 per cent; SP-4, 17.6 per cent. In the parotid saliva of rats fed the liquid diet, proteins corresponding to Peak A and SP-2 were not present, the proportion of BPRP in SP-4 was increased to almost 90 per cent while the proportions of material in SP-1 and SP-3 were reduced to 3 and 8 per cent, respectively. In the saliva of rats subjected to chronic isoproterenol treatment, a protein corresponding to SP-4 was not present; proteins corresponding to Peak A, SP-1 and SP-3 were present and in amounts similar to their proportion in untreated rats although material in SP-2 increased to 36 per cent.     

Fractionation of salivary micelle-like structures by gel chromatography

Globular structures have been demonstrated in human parotid saliva by transmission electron microscopy and photon correlation spectroscopy. The aim of this study was to fractionate these salivary globular structures for analytical and preparative purposes using a gel-filtration material capable of separating spherical particles up to 300-400 nm in diameter. Freshly obtained parotid saliva was applied to a Sephacryl S-1000 column. Peak fractions were collected and prepared for transmission electron microscopy (TEM) or for amino acid analysis. Bovine milk was included as the casein micelles by TEM appear to be similar to the salivary aggregates and their elution profiles are known. The salivary globular structures were eluted in one major peak. TEM of negatively stained samples from the peak fractions demonstrated globular protein aggregates consistent with the salivary structures in parotid saliva. Amino acid analysis showed characteristic amino acid profiles with unusual high levels of proline, 40-45%. The casein micelles were eluted in one major peak and separated from the whey proteins. This study indicates that the salivary globular structures can be isolated by gel chromatography. The amino acid analysis indicates that proline-rich proteins may be an important fraction of the salivary globular structures.     

Parotid salivary basic proline-rich proteins inhibit HIV-I infectivity

OBJECTIVE: The objective of this study was to investigate the molecular nature, spectrum of activity and mechanism(s) of action of those human parotid basic proline-rich proteins that exhibit anti-HIV-I activity. DESIGN: Fractions containing the basic proline-rich proteins were obtained from human parotid saliva of presumed HIV-I non-infected human subjects and characterized with respect to their purity, apparent molecular size and their ability to inhibit the infectivity of T-tropic and M-tropic strains of HIV-I. SUBJECTS, MATERIALS AND METHODS: Stimulated parotid saliva samples were collected from human subjects who denied having any risk factors for HIV-I infection and whose parotid salivas inhibited HIV-I infectivity. Such samples were subjected to affinity, molecular sieve and ion exchange chromatography to isolate individual salivary components. Those fractions demonstrating anti-HIV-I activity were analyzed by SDS-PAGE in order to assess their purity and determine their apparent molecular weights. HIV-I inhibitory activity was determined using HIV-I strains LAI and BaL in a Hela cell-derived multinuclear activation of a galactosidase indicator (MAGI) assay. Amino acid analyses were performed on some fractions. RESULTS: Recombinant gp120-CH-Sepharose chromatography of one subject's parotid saliva revealed specific binding of human parotid basic proline-rich proteins, most prominently one with an apparent molecular weight of 37 kDa. Molecular sieve and cation exchange chromatography yielded a fraction greatly enriched in this protein which amino acid analysis confirmed was proline-rich. A similar fraction from two other subjects also contained basic proline-rich proteins of similar molecular size. These fractions inhibited both T-tropic and M-tropic strains of HIV-I when assayed in the MAGI system. Since SLPI activity is not observable in the MAGI assay, this inhibition was not due to SLPI. The presence of thrombospondin-I (TSP-I) in the active fractions was precluded on the basis of SDS-PAGE examination. CONCLUSIONS: Specific basic proline-rich proteins in human parotid saliva possess significant anti-HIV-I activity independent of that attributable to SLPI or TSP-I. Since the inhibition is detectable with the MAGI assay, its mechanism of action involves virus-host cell interaction prior to the introduction of the tat gene product into the host cell and may be through the binding of the basic proline-rich proteins to the HIV-I gp120 coat of the virus.     

Chronic treatment with beta adrenergic agonists and antagonists alters the composition of proteins in rat parotid saliva

This investigation was undertaken to determine the role of beta-adrenergic receptors in the regulation of the protein composition of rat parotid saliva. Chronic treatment of rats with dobutamine, a beta 1-adrenergic agonist, resulted in changes in parotid saliva volume, protein concentration, and composition which were essentially the same as those changes which occurred following chronic treatment with isoproterenol, a non-specific beta-adrenergic agonist. Chronic treatment with the beta 2-adrenergic agonist, terbutaline, had no effect on parotid saliva volume, protein concentration, or composition. Chronic treatment of rats with a beta 1-adrenergic antagonist, metoprolol, had different effects on saliva dependent on the manner by which the drug was delivered. Twice-daily injections of metoprolol led to a decrease in flow rate, but protein concentration and composition were unaltered. When metoprolol was delivered by surgically implanted osmotic minipumps, neither the flow of parotid saliva nor its concentration of protein was altered; however, there was a reduction in the proportion of proline-rich proteins in saliva. Comparable changes in parotid saliva protein composition occurred when the minipumps delivered propranolol, a non-specific beta-adrenergic antagonist. Chronic treatment of rats with an alpha 2-adrenergic agonist (clonidine) or antagonist (yohimbine) was without effect on parotid saliva flow rate, protein concentration, or composition. These findings suggest that the synthesis of proline-rich proteins is regulated, in part, by beta-adrenergic receptor stimulation, and primarily by the beta 1-receptor subtype.     

Parotid secretory granules: crossroads of secretory pathways and protein storage

Saliva plays an important role in digestion, host defense, and lubrication. The parotid gland contributes a variety of secretory proteins-including amylase, proline-rich proteins, and parotid secretory protein (PSP)-to these functions. The regulated secretion of salivary proteins ensures the availability of the correct mix of salivary proteins when needed. In addition, the major salivary glands are targets for gene therapy protocols aimed at targeting therapeutic proteins either to the oral cavity or to circulation. To be successful, such protocols must be based on a solid understanding of protein trafficking in salivary gland cells. In this paper, model systems available to study the secretion of salivary proteins are reviewed. Parotid secretory proteins are stored in large dense-core secretory granules that undergo stimulated secretion in response to extracellular stimulation. Secretory proteins that are not stored in large secretory granules are secreted by either the minor regulated secretory pathway, constitutive secretory pathways (apical or basolateral), or the constitutive-like secretory pathway. It is proposed that the maturing secretory granules act as a distribution center for secretory proteins in salivary acinar cells. Protein distribution or sorting is thought to involve their selective retention during secretory granule maturation. Unlike regulated secretory proteins in other cell types, salivary proteins do not exhibit calcium-induced aggregation. Instead, sulfated proteoglycans play a role in the storage of secretory proteins in parotid acinar cells. This work suggests that unique sorting and retention mechanisms are responsible for the distribution of secretory proteins to different secretory pathways from the maturing secretory granules in parotid acinar cells.     

Correlations between human salivary levels of lysozyme, lactoferrin, salivary peroxidase and secretory immunoglobulin A with different stimulatory states and over time

Within-subject correlations for the levels of these salivary proteins were determined in unstimulated and stimulated parotid saliva collected from 8 subjects and for stimulated parotid saliva collected from the same subjects once a week for a 4-week period. Initial correlations between unstimulated and stimulated samples were high and statistically significant (p less than 0.05) for all four proteins. When data were adjusted for variation attributable to flow rate and total protein, some correlations remained the same and those for lysozyme, lactoferrin and salivary peroxidase increased. However, the correlation for secretory immunoglobulin A decreased to a point where it was no longer statistically significant. In the weekly comparison, within-subject correlations across weeks were significant (p less than or equal to 0.05) for lysozyme, lactoferrin and salivary peroxidase, but not for immunoglobulin A. After adjustment for flow rate and total protein, the pattern of correlation was unchanged. Thus the relative rankings of subjects for levels of lysozyme, lactoferrin or salivary peroxidase may be consistent across stimulatory states, even though absolute concentrations may change; levels of these proteins in stimulated parotid saliva may be maintained over time. Secretory immunoglobulin A appears to be more subject to short-term variation.     

Demonstration of proline-rich proteins in rabbit parotid saliva and partial characterization of some of the proteins

Rabbit parotid saliva was collected by cannulation of the secretory duct in anaesthetized animals. Proteins which cross-react with antibodies to human acidic proline-rich proteins were demonstrated in the secretion and fractionated by chromatography on an immunosorbent and CM32-cellulose. At least 5 proline-rich proteins were identified with molecular weights ranging from 19,000 to 61,000 as determined by sodium dodecylsulphate acrylamide gel electrophoresis. The major immunoreactive components were basic proteins with similar size and charge properties. In two of the proteins, proline, glycine and glutamic acid or glutamine accounted for 84 or 99 per cent of all residues. In contrast to proline-rich proteins from other species, proline constituted only 13 or 17 per cent of total amino acids.     

Effects of a beta-blocking agent, timolol maleate, on saliva in healthy volunteers

The effects of timolol maleate on the secretion and composition of human saliva were studied in vivo. Eight healthy volunteers received orally 10 mg timolol maleate. Stimulated parotid saliva samples, resting whole saliva samples, and blood samples were collected immediately before and four times after the drug intake at intervals of 1 h. The levels of total protein, lysozyme, IgA, IgG and IgM, salivary peroxidase, myeloperoxidase, lactoferrin, amylase, thiocyanate (SCN-), and hypothiocyanite (OSCN-) were analyzed from saliva samples. Drug levels were measured both from parotid saliva and blood samples. Results were compared to the analyses of the samples collected in a similar way but without administration of any drugs. Decreased levels of total protein, lactoferrin, amylase, and salivary peroxidase were observed in parotid saliva after a single oral dose of timolol maleate. No such decrease was found in lysozyme, myeloperoxidase, SCN-, OSCN-, or immunoglobulins. Salivary flow rate was not significantly changed after drug intake. The results suggest that the beta-blocking drug may cause qualitative changes in the composition of saliva by inhibiting the synthesis and/or release of acinar proteins.     

Isolation and characterization of six proteins from rabbit parotid saliva belonging to a unique family of proline-rich proteins

Proline-rich proteins are major components of salivary secretion from humans non-human primates, rats, hamsters and rabbits. They are also synthesized in mice in response to chronic stimulation by beta agonists. This study to provide an understanding of the structural and genetic relationships within these families of proteins to determine the possible function of the proline-rich proteins. Rabbit parotid saliva was collected and proline-rich proteins were affinity purified using goat antibodies to human proline-rich proteins. Purification was achieved by repeated cation exchange chromatography on a Mono S column a Fast Protein Liquid Chromatography system. Six basic proline-rich proteins were purified. The apparent molecular weights were between 75,000 and 125,000, based on their mobilities in sodium dodecyl sulphate-polyacrylamide gel electrophoresis. Glycine, glutamine (and glutamate) and proline accounted for 79-87% of total amino acids in all proteins, but proline was present in smaller amounts (17-21%) than in proline-rich proteins from other species. All proteins were glycosylated but not phosphorylated. Circular dichroism of two proline-rich proteins, MS7A and MS5B, indicated the absence of secondary structure. The N-terminal sequences of three proteins electro-eluted after preparative gel electrophoresis were determined. A high degree of similarity was found in various regions of mouse, rat, monkey and human proline-rich proteins. Rabbits thus synthesize constitutively a family of proteins that are immununologically and structurally related to proline-rich proteins other species.