Cuticular proteins from the blue crab alter in vitro calcium carbonate mineralization

Coblentz, Francie E; Shafer, Thomas H.; Roer, Robert D.

doi:10.1016/s0305-0491(98)10117-7

Cited by 43 publications

(48 citation statements)

References 22 publications

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“…This evidence includes the existence of a highly soluble initial phase, especially in the IPS, which is consistent with reports of similar transient amorphous forms in sea urchin spicules (Beniash et al, 1997) and larval mollusc shells (Weiss et al, 2002), and more stable forms of amorphous calcium carbonate in ascidian spicules (Aizenberg et al, 2002) and the calcium storing organ of an isopod (Raz et al, 2003). Therefore, the glycoprotein(s) described in this research and predicted to inhibit calcite nucleation here and in the past (Shafer et al, 1995;Coblentz et al, 1998) may actually inhibit the initial deposition of amorphous calcium carbonate.…”

Section: Discussionsupporting

confidence: 91%

“…Since the IPS are the first regions to calcify in the exocuticle (GiraudGuille, 1984;Dillaman et al, 2001;Hequembourg, 2002), the 3·h post-molt antigen localization represents the negative image of the calcification pattern that will appear somewhat later. This finding adds a spatial correlation to the temporal correlation we have described between changes in these glycoprotein bands and mineralization (Shafer et al, 1995), and strengthens our hypothesis that the unaltered glycoproteins represent inhibitors of crystal nucleation (Coblentz et al, 1998). Their early post-molt alteration in the IPS, but not the prisms, could control the pattern of mineralization in the exocuticle.…”

Section: Discussionsupporting

confidence: 87%

“…The C. sapidus glycoprotein material previously studied (Shafer et al, 1994(Shafer et al, , 1995Coblentz et al, 1998) was purified from total soluble protein extracts of cuticle from crabs killed at ecdysis. A jacalin lectin affinity column was used to separate glycoproteins with O-linked carbohydrates, which were then separated on the basis of size using a gel filtration column (Fig.·1).…”

Section: Discussionmentioning

confidence: 99%

“…It is unclear whether proteolysis causes the disappearance of these proteins or if deglycosylation causes the loss of lectin and PAS affinity. Subsequently, Coblentz et al (1998) showed that two heavily glycosylated proteins identical in apparent molecular mass to the previously described proteins associate with CaCO3 crystals in a solutionbased mineralization assay, but decrease their affinity for the crystals as time increases post-molt. A model in which these glycoproteins act as pre-molt inhibitors of calcification in the exocuticle was proposed (Coblentz et al, 1998).…”

mentioning

confidence: 90%

“…Subsequently, Coblentz et al (1998) showed that two heavily glycosylated proteins identical in apparent molecular mass to the previously described proteins associate with CaCO3 crystals in a solutionbased mineralization assay, but decrease their affinity for the crystals as time increases post-molt. A model in which these glycoproteins act as pre-molt inhibitors of calcification in the exocuticle was proposed (Coblentz et al, 1998). The proteins are hypothesized to interact with sites of nucleation on the chitin-protein fibers, thereby limiting crystal formation despite supersaturating concentrations of calcium and bicarbonate.…”

mentioning

confidence: 90%

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Purification of a soluble glycoprotein from the uncalcified ecdysial cuticle of the blue crabCallinectes sapidusand its possible role in initial mineralization

Tweedie

Coblentz

Shafer

2004

Journal of Experimental Biology

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A heavily glycosylated soluble protein was purified using a combination of lectin affinity and size exclusion chromatography from a soluble extract of uncalcified dorsal cuticle of blue crab Callinectes sapidus removed at ecdysis. Similarities in apparent molecular mass and carbohydrate composition suggest that this protein is the same species previously shown to disappear from soluble extracts coincidentally with the onset of mineral deposition in the newly exposed post-molt cuticle. The amino acid sequence of the N-terminal portion of the core polypeptide was determined and polyclonal antibodies were raised against both the purified glycoprotein and the peptide. Immunoblots of unfractionated soluble extracts taken at various times post-molt illustrated that the antipeptide antibody recognized several polypeptides with electrophoretic mobilities that differ from the purified glycoprotein. These bands may be deglycosylation products which would not have been purified due to different lectin affinity or size. Immunohistochemical analysis indicated uniform protein distribution in the exocuticle at ecdysis, but decreased antibody binding at the interprismatic septa by 2·h post-molt. The location of the protein is therefore the negative image of the calcification pattern in the exocuticle and provides a spatial pattern to correlate with the previously reported temporal events. This strengthens the hypothesis that the glycoprotein under investigation is an inhibitor of calcite nucleation or of initial amorphous calcium carbonate accumulation.

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Section: Discussionsupporting

confidence: 91%

Section: Discussionsupporting

confidence: 87%

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 90%

mentioning

confidence: 90%

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Purification of a soluble glycoprotein from the uncalcified ecdysial cuticle of the blue crabCallinectes sapidusand its possible role in initial mineralization

Tweedie

Coblentz

Shafer

2004

Journal of Experimental Biology

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Immunocytochemical localization of actin and tubulin in the integument of land crab (Gecarcinus lateralis) and lobster (Homarus americanus)

2000

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The crustacean integument consists of the exoskeleton and underlying epithelium and associated tissues. The epithelium, which is composed of a single layer of cells, is responsible for the cyclical breakdown and synthesis of the exoskeleton associated with molting (ecdysis). During premolt (proecdysis) the epithelial cells lengthen and secrete the two outermost layers (epicuticle and exocuticle) of the new exoskeleton while partially degrading the two innermost layers (endocuticle and membranous layer) of the overlying old exoskeleton. This increased cellular activity is associated with increased protein synthesis and a change in cell shape from cuboidal to columnar. The cytoskeleton, composed of microfilaments (actin) and microtubules (tubulin), plays important roles in the intracellular organization and motility of eukaryotic cells. Immunoblot analysis shows that the land crab exoskeleton contains actin, tubulin, and actin‐related proteins (Varadaraj et al. 1996. Gene 171:177–184). In the present study, immunocytochemistry of land crab and lobster integument showed that both proteins were localized in various cell types, including epithelia, connective tissue, tendinal cells, and blood vessels. Muscle immunostained for actin and myosin, but not for tubulin. The membranous layer of land crab (the other layers of the exoskeleton were not examined) and membranous layer and endocuticle of lobster also reacted specifically with anti‐β‐actin and anti‐α‐tubulin monoclonal antibodies, but not with an anti‐myosin heavy chain antibody. During proecdysis immunolabeling of the membranous layer decreased probably due to protein degradation. The staining intensity for actin and tubulin in the proecdysial epithelium was similar to that in the intermolt (anecdysial) epithelium, suggesting that there was a net accumulation of both proteins proportional to the increase in cellular volume. These results support the previous biochemical analyses and, more specifically, localize actin and tubulin in exoskeletal structures, suggesting that they may serve both intracellular and extracellular functions in crustaceans. J. Exp. Zool. 286:329–342, 2000. © 2000 Wiley‐Liss, Inc.

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Biomimetic Lessons for Processing Chitin‐Based Composites

Wilson¹,

Omokanwaye²

2013

Biopolymer Nanocomposites

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Cuticular proteins from the blue crab alter in vitro calcium carbonate mineralization

Cited by 43 publications

References 22 publications

Purification of a soluble glycoprotein from the uncalcified ecdysial cuticle of the blue crabCallinectes sapidusand its possible role in initial mineralization

Purification of a soluble glycoprotein from the uncalcified ecdysial cuticle of the blue crabCallinectes sapidusand its possible role in initial mineralization

Immunocytochemical localization of actin and tubulin in the integument of land crab (Gecarcinus lateralis) and lobster (Homarus americanus)

Biomimetic Lessons for Processing Chitin‐Based Composites

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