Cleavage of Rabbit Myelin Basic Protein by Thrombin

Law, Mona J.; Martenson, Russell E.; Deibler, Gladys E.

doi:10.1111/j.1471-4159.1984.tb02714.x

Cited by 35 publications

(19 citation statements)

References 34 publications

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“…Actin [47] Actin [47] Actin [47] Ovine growth hormone [49] Bovine growth hormone [49] Human growth hormone [49] Human chorionic gonadotropin [54] Rabbit myelin basic protein [55] Rabbit myelin basic protein [55] Tubulin [56] Dynorphin Somatostatin-28'…”

Section: The Rather Large Hydrophobic Pocket Of A-thrombin Enhances Imentioning

confidence: 99%

“…Eel calcitoninb PTH fragment Antibody K-chain [59] Antibody i-chain [59] Antibody i-chain [59] Antibody A-chain [59] Antibody A-chain [59] P [54] Human factor XI11 [57] Human factor XI11 [57] Gastrin-releasing peptide Bovine &-thrombin [33] Antithrombin TIT [52] Rabbit myelin basic protein [55] Rabbit myelin basic protein [55] Cholecystokinin [26] Antibody i-chain [59] Antibody L-chain [59] Secretin [25] Myoglobin [48] Chymotrypsinogen A [48] Trysinogen [48] Human m-thrombin [34] Troponin [51] Rabbit myelin basic protein [55] Cholecystokinin their optimal P2-P4 structures. This effect is best demonstrated by the significant difference of thrombin susceptibility between salmon and eel calcitonins.…”

Section: Salmon Calcitoninbmentioning

confidence: 99%

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Thrombin specificity

Chang

1985

European Journal of Biochemistry

202

108

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a-Thrombin cleavage of 30 polypeptide hormones and their derivatives were analysed by quantitative aminoterminal analysis. The polypeptides included secretin, vasoactive intestinal polypeptide, cholecystokinin fragment, dynorphin A, somatostatins, gastrin-releasing peptide, calcitonins and human parathyroid hormone fragment. Most of them were selected mainly on the ground that they contain sequence structures homologous to the well known tripeptide substrates of a-thrombin. All selected polypeptides have one single major cleavage site and both Arg-Xaa and Lys-Xaa bonds were found to be selectively cleaved by a-thrombin. Under fixed conditions (1 nmol polypeptide/0.5 NIH unit a-thrombin in 20 pl of 50 mM ammonium bicarbonate at 25 "C), the time required for 50% cleavage ranges from less than 1 min to longer than 24 h. Heparin invariably enhanced thrombin cleavage on all polypeptide analysed. The optimum cleavage site for a-thrombin has the structures of (a) P4-P3-Pro-Arg-P 1'-P2', where P3 and P4 are hydrophobic amino acid and PI', P2' are nonacidic amino acids and (b) P2-Arg-P l', where P 2 or P 1' are Gly. The requirement for hydrophobic P 3 and P4 was further demonstrated by the drastic decrease of thrombin cleavage rates in both gastrin-releasing peptide and calcitonins after chemical removal of hydrophobic P 3 and P4 residues. The requirement for nonacidic P1' and P2' residues was demonstrated by the drastic increase of thrombin cleavage rates in both calcitonin and parathyroid hormone fragments, after specific chemical modification of acidic P 1' and P2' residues. These findings confirm the importance of hydrophobic P2-P4 residues for thrombin specificity and provide new evidence to indicate that apolar P1' and P2' residues are also crucial for thrombin specificity. It is concluded that specific cleavage of polypeptides by a-thrombin can be SedSOnably predicted and that chemical modification can be a useful tool in enhancing thrombin cleavage. a-Thrombin is a trypsin-like serine proteinase which preferentially cleaves polypeptide substrates at Arg/Lys-Xaa bonds [I -31. Unlike trypsin, a-thrombin displays highly restricted specificity toward protein substrates. This specificity is best known by its selective cleavage of two Arg-Gly bonds from amongst 181 Arg/Lys-Xaa bonds in fibrinogen [4]. Attempt to delineate the structural element in fibrinogen which accounts for restricted thrombin cleavage has mostly focussed on the fibrinogen Aa chain [5 -81 and has led to the findings that (a) the Aa (1-23) fragment contains a fundamental structure element for thrombin binding Table 1 for numbering of residues.) The recognization of the importance of hydrophobic P2 and P9 residues has further led to the development of a series of useful tripeptide substrates [9 -131 and inhibitors [I 4, 151. Another source of understanding of thrombin specificity comes from thrombin cleavage at non-fibrinogen polypeptides. A number of peptides and proteins have been Abbreviations. DABS-GI, dimethylaminoazobenzene sulfonyl chloride;...

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Section: The Rather Large Hydrophobic Pocket Of A-thrombin Enhances Imentioning

confidence: 99%

Section: Salmon Calcitoninbmentioning

confidence: 99%

Thrombin specificity

Chang

1985

European Journal of Biochemistry

202

108

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“…Peptide Cl vs. peptide C2 at the same concentration. the basis of the results of Law et al [1984] for rabbit MBP, it can be assumed that cleavage took place be tween Arg(96) and Thr(97) resulting in Tl 1-96 and T2 97-169. It is of interest that during purification peptide T2.…”

Section: R -And Vr-potentials Glutamatementioning

confidence: 99%

“…BMBP (peak 2) was cleaved by thrombin as described by Law et al [1984] for single site cleavage of rabbit MBP, with slight modifi cations. The reaction was carried out in a mixture (pH 7.4) of ammonium hydrogen carbonate (0.5 M ) and ammonium acetate (0.1 M ) instead of an imidazole buffer, enabling buffer removal by simple lyophilization.…”

Section: Cleavage O F Peak 2 By Thrombin and Isolation O F The Peptidesmentioning

confidence: 99%

“…Plasmin rapidly degrades MBP in isolated central nervous system (CNS) myelin in vitro [Cammer et al, 1978;Inuzuka et al, 1984], Cathepsin D is increased during inflammatory demyelination of the CNS, for instance in brains of animals with experimental allergic encephalomyelitis [Rauch et al, 1973], This sug gests that it is involved in the normal metabolism and pathological degradation of CNS proteins. Thrombin is an enzyme known to cleave specifically at the carboxyl peptide bond of certain arginine residues in a variety of proteins [Blombaeck et al, 1977], Law et al [ 1984] have shown that it is essentially possible to cleave rabbit MBP completely and specifically at a single site, the Arg(95)-Thr(96) bond.…”

Section: Introductionmentioning

confidence: 99%

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Effects of Peptides from Bovine Myelin Basic Protein on the Bioelectrical Activity of the Frog Spinal Cord

Brugger¹,

Honegger²,

Kaeser³

1988

Eur Neurol

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Both bovine and human myelin basic protein (MBP) have been shown to have electrophysiological activity. As MBP is susceptible to proteolytic degradation, our aim was to discover whether the resulting peptides retained this activity. Bovine MBP was completely cleaved by plasmin into at least nine peptides. The electrophysiological activities of this peptide mixture and of bovine MBP were directly compared on the hemisected frog spinal cord. The peptide mixture and intact bovine MBP had quantitatively and qualitatively similar effects (dose-dependent long-lasting depolarization, about 100 times more active than glutamate). Four peptides (molecular weights 14,000, 10,500, 8,000, 4,500) from thrombin or cathepsin D cleavage of bovine MBP also showed electrophysiological activity, positively correlated to their molecular weights. As MBP-like material occurs in increased concentrations in the cerebrospinal fluid during demyelinating diseases, peptides resulting from proteolytic degradation of MBP, e.g. in demyelinating foci of multiple sclerosis, might cause neuronal disturbances.

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Myelin basic protein‐specific T‐cell responses in identical twins discordant or concordant for multiple sclerosis

Martin

Voskuhl

Flerlage

et al. 1993

Annals of Neurology

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Although multiple sclerosis (MS) is thought to be an autoimmune disease, the target antigen of the immune response is unknown. Both myelin basic protein (MBP) and proteolipid protein (PLP) have been considered candidate autoantigens. Because the immune response to either foreign or self antigens is influenced by the genetic background of the host, the importance of these candidate antigens has been difficult to establish in humans because of genetic diversity. To eliminate genetic differences in MS patients and healthy controls, we have studied the MBP-specific T-cell response in 6 sets of identical twins, 3 of which were concordant and 3 discordant for MS. A total of 638 short-term T-cell lines were established and characterized for MBP-specific proliferative and cytotoxic activity, fine specificity, and human leukocyte antigen (HLA) restriction. Similar frequencies of MBP-specific T cells were observed in affected and unaffected individuals. A slightly higher percentage of cytotoxic T-cell lines was found in affected individuals. For most of the cell lines, the restriction elements were the HLA class II antigens that have been reported previously to be associated with MS; no important differences with respect to HLA restriction were found between the patients and healthy individuals. The peptide epitopes of MBP that were recognized most frequently by the T-cell lines were those previously shown to be immunodominant. Differences in specificity were seen in some discordant twins indicating that, despite genetic identity, the MBP-specific T-cell repertoire may be shaped differently. These findings indicate that differences in frequency, peptide specificity, or HLA restriction are not sufficient to implicate MBP-specific T cells in the pathogenesis of MS. However, the T-cell response to MBP may still represent one necessary component with disease occurring when this response is combined with other host characteristics such as regulation of cytokine-, adhesion molecule-, or HLA-antigen expression in the nervous system or immunoregulatory mechanisms.

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Cleavage of Rabbit Myelin Basic Protein by Thrombin

Cited by 35 publications

References 34 publications

Thrombin specificity

Thrombin specificity

Effects of Peptides from Bovine Myelin Basic Protein on the Bioelectrical Activity of the Frog Spinal Cord

Myelin basic protein‐specific T‐cell responses in identical twins discordant or concordant for multiple sclerosis

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