The transpeptidase system which crosslinks fibrin by γ-glutamyl-ε-lysine bonds

Lóránd, L.; Downey, Jeanne E.; Gotoh, Toshio; Jacobsen, Ann-Christin; Tokura, Seiichi

doi:10.1016/0006-291x(68)90734-1

Cited by 160 publications

(53 citation statements)

References 15 publications

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“…4, b) (24,26,27) and LTG (25), especially around their active sites. XIIIa, the final component in the coagulation pathway, plays an important role in the stabilization of fibrin clots by covalently crosslinking fibrin monomers through y-glutamyl-E-lysine bridges and by preventing proteolysis (35). Liver transglutaminase has some ofthe activity associated with the plasma membrane and may be responsible for forming covalently crosslinked matrices of proteins at sites of cell-to-cell contact (36).…”

Section: Methodsmentioning

confidence: 99%

Molecular cloning of human protein 4.2: a major component of the erythrocyte membrane.

Sung¹,

Chien²,

Chang³

et al. 1990

Proc. Natl. Acad. Sci. U.S.A.

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Protein 4.2 (P4.2) comprises -5% of the protein mass of human erythrocyte (RBC) membranes. Anemia occurs in patients with RBCs deficient in P4.2, suggesting a role for this protein in maintaining RBC stability and integrity. We now report the molecular cloning and characterization of human RBC P4.2 cDNAs. By immunoscreening a human reticulocyte cDNA library and by using the polymerase chain reaction, two cDNA sequences of 2.4 and 2.5 kilobases (kb) were obtained. These cDNAs differ only by a 90-base-pair insert in the longer isoform located three codons downstream from the putative initiation site. The 2.4-and 2.5-kb cDNAs predict proteins of -77 and =80 kDa, respectively, and the authenticity was confimed by sequence identity with 46 amino acids of three cyanogen bromide-cleaved peptides of P4. Subcloning and Sequence Analysis. cDNA inserts from positive phage clones were subcloned into pBS(+) plasmids (Stratagene). Unidirectional deletion clones were generated by using BAL-31 exonuclease (13), and cDNA fragments were sequenced with T3 and T7 primers by the dideoxynucleotide chain-termination method (14). Sequence analysis and GenBank data base searches were performed by IBI Pustell sequence analysis software (International Biotechnologies). 5'-End Extension of cDNA. Three oligonucleotides were prepared in a technique based on the polymerase chain reaction (PCR) to synthesize the missing 5' sequence of the partial cDNA clone: pl was composed of nucleotides (nt) 7-23 of clone 7 (c.7); p2 was complementary to nt 36-52 of c.7 plus an EcoRI restriction site at its 5' end; p3 was composed of the EcoRI polylinker and the poly(dC) originally used for the first-strand cDNA synthesis when the library was constructed. The sequences of pl, p2, and p3 were, respectively, 5'-dTGAGGATGCTGTGTTCC-3', 5'-dTCGAAlJCGTACTC-CATGCGCTGAG-3', and 5'-dGCGGLAAlTCCCCCCCCCC-CCCC-3', with the EcoRI sites underlined. p2 and p3 were used as PCR primers, and the reticulocyte cDNA library (5 ,u with 106 phages per pl) was used as a template. The reaction product was electrophoresed and stained with ethidium bromide. The major band was excised and subcloned into pGEM 3zf plasmids (Promega). From >500 transformants, 24 colonies were randomly chosen to make minipreparations of plasmid DNA, of which 80%o were positive when hybridized with 32P-labeled pl. tTo whom reprint requests should be addressed. IThe sequences reported in this paper have been deposited in the GenBank data base (accession nos. M30646 and M30647). 955The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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Section: Methodsmentioning

confidence: 99%

Molecular cloning of human protein 4.2: a major component of the erythrocyte membrane.

Sung¹,

Chien²,

Chang³

et al. 1990

Proc. Natl. Acad. Sci. U.S.A.

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“…2 for review). The second is the e(-y-glutamyl)lysine amide bonds formed during the stabilization of fibrin clot (3)(4)(5), and reported to occur in cultures of chicken-breast muscle (6), I-cell membrane (6), native wool keratin (7), and the citrulline-containing protein fraction of hair (8).…”

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confidence: 99%

Transglutaminase from Hair Follicle of Guinea Pig

Chung

Folk

1972

Proc. Natl. Acad. Sci. U.S.A.

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Two transglutaminases are found in homogenates of the inner root sheaths of guinea pig hairfollicles. One is indistinguishable from the well-characterized liver transglutaminase [J. Biol. Chem., 246, 1093]. The other, which is present in far greater quantity, has not been detected in other organs or tissues. Gel filtration and polyacrylamide gel electrophoresis studies indicate that the native hair-follicle enzyme, of molecular weight 54,000, is composed of two subunits of identical molecular weight. Specificity studies suggest that the intermolecular cross-linking of fibrin and fibrinogen that is catalyzed by this enzyme is a result of the formation of e(y-glutamyl)lysine bonds. The probable participation of hair-follicle transglutaminase in the formation of these cross-links in the proteins of hair is discussed.The essential role in protein structure of intramolecular covalent crosslinks, in the form of disulfide bonds, has been recognized for many years (ref. 1 for review). There is an increasing awareness of the physiological importance of other covalent crosslinks, within and between protein molecules. Two types of chemical bonds are known to comprise these crosslinks. One, which occurs in collagen and elastin, is the combination of lysine-and hydroxylysine-derived aldehyde condensation products and Shiff bases formed by coupling of these aldehydes with e-amino groups (ref. 2 for review). The second is the e(-y-glutamyl)lysine amide bonds formed during the stabilization of fibrin clot (3-5), and reported to occur in cultures of chicken-breast muscle (6), I-cell membrane (6), native wool keratin (7), and the citrulline-containing protein fraction of hair (8).Work in this laboratory has centered on guinea pig liver transglutaminase, an enzyme that catalyzes formation of -y-glutamyl amide bonds (9, 10) by way of an acyl-transfer mechanism (I1). The wide occurrence and obvious importance of y-glutamyl amide bonds has led us to focus attention on the various enzymes responsible for their formation. Transglutaminases derived from plasma and platelet protransglutaminases (blood coagulation Factor XIII) through thrombin activation (ref. 12 for review) are responsible for the stabilization of fibrin clot, a consequence of formation of e(yglutamyl)lysine bridges between fibrin monomers. Purified transglutaminase from guinea pig liver stabilizes both fibrin (13) and fibrinogen (14) The other, which is present in far greater quantity, is distinctly different from previously described enzymes, and is the principal subject of this paper. MATERIALS AND METHODSSkins from 20-to 25-week-old guinea pigs were obtained packed in dry ice from Rockland (Gilbertsville, Pa. RESULTS Isolation of enzymesStep 1. Hair follicles of the guinea pig were,exposed by the wax-sheet method (17, 18) with a beeswax-rosin mixture at a temperature not exceeding 55°C. The inner-root sheaths were

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“…In the sixties there were two important developments. First, the chemical nature of the cross-link formed in fibrin (Laki and Lorand, Science 1948) by blood coagulation factor XIIIa, a plasma transglutaminase (Loewy 1968), was revealed (Pisano et al 1968;Matatic and Loewy 1968;Lorand et al 1968). Second, the purification of tissue transglutaminase (later named transglutaminase 2; TG2) from guinea pig liver could be achieved (Folk and Cole 1966) in the NIH laboratory of Jack Folk, opening the way to the characterization of its enzymatic properties in details; the findings were summarized in the early seventies by Folk and Chung providing a basis of further developments in the field (Folk and Chung 1973).…”

Section: Introductionmentioning

confidence: 99%

Polymorphism of transglutaminase 2: unusually low frequency of genomic variants with deficient functions

2011

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Transglutaminase 2 (TG2) is a multifunctional member of an enzyme family: it modifies glutamine residues by cross-linking proteins and incorporating primary amines into them, has protein disulphide isomerase and protein kinase activities, mediates trans-membrane signal transduction and interactions between cell surface proteins and the extracellular matrix. These unusual multiple roles encoded into one polypeptide chain suggest that genomic variations in the TGM2 gene should be limited. Indeed, the available information in databases shows that unlike in the case of most other transglutaminases there are no common single nucleotide polymorphisms in exons of human TGM2. We collected data on and produced some of the rare genetic variants of TGM2 by site directed mutagenesis and found that some were less stable than the most abundant (wild type) enzyme variant and the majority had deficient transamidating activity. Further studies are required to clarify the pathologic significance of these rare TGM2 alleles in the human population.

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The transpeptidase system which crosslinks fibrin by γ-glutamyl-ε-lysine bonds

Cited by 160 publications

References 15 publications

Molecular cloning of human protein 4.2: a major component of the erythrocyte membrane.

Molecular cloning of human protein 4.2: a major component of the erythrocyte membrane.

Transglutaminase from Hair Follicle of Guinea Pig

Polymorphism of transglutaminase 2: unusually low frequency of genomic variants with deficient functions

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