Measurement of bone resorption in serum using antibodies reactive with an isomerized form of an 8 amino acid sequence of the C-telopeptide of type I collagen

1996 ◽  
Vol 6 (S1) ◽  
pp. 193-193
Author(s):  
M. Bonde ◽  
P. Garnero ◽  
C. Fledelius ◽  
S. Christgau ◽  
P. Qvist ◽  
...  
Keyword(s):  
Amino Acid ◽  
Bone Resorption ◽  
Amino Acid Sequence ◽  
Type I Collagen ◽  
Type I

scholarly journals Extracellular-matrix degradation at acid pH. Avian osteoclast acid collagenase isolation and characterization

1993 ◽  
Vol 290 (3) ◽  
pp. 873-884 ◽  
Author(s):  
H C Blair ◽  
S L Teitelbaum ◽  
L E Grosso ◽  
D L Lacey ◽  
H L Tan ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Substrate Specificity ◽  
Cathepsin B ◽  
Type I Collagen ◽  
Sequence Similarity ◽  
Bone Matrix ◽  
Collagenolytic Activity ◽  
Type I ◽  
Isolation And Characterization

Osteoclasts degrade bone matrix, which is mainly type I collagen and hydroxyapatite, in an acidic extracellular compartment. Thus we reasoned that osteoclasts must produce an acid collagenase. We purified this enzyme, a 31 kDa protein, from avian osteoclast lysates (in 100 mM acetate/1 mM CHAPS/1 mM dithiothreitol, pH 4.4), fractionated by (NH2)2SO4 precipitation, gelatin-affinity, cation exchange, and gel filtration. Fraction activity was measured using diazotized collagen or 3H-labelled cross-linked collagen (decalcified and trypsin-treated metabolically L-[4,5-3H]proline-labelled bone) as substrates. Iodoacetate, leupeptin, antipain, pepstatin and mercurials inhibited collagenolysis by the isolated proteinase; mercurial derivatives could not be re-activated by dithiothreitol. Collagen degradation was maximal at pH 4.4; purified proteinase reproduced the collagenolytic activity of cell lysates. The N-terminal amino acid sequence from the isolated protein and its CNBr degradation fragments showed sequence similarity to mammalian cathepsin Bs, and near-identity with avian liver cathepsin B. Peptide substrate specificity of the osteoclastic enzyme resembled those of mammalian cathepsin B and its avian liver counterpart, but degradation of low-molecular-mass substrates by the osteoclastic enzyme was slower, reflecting generally lower kcat. values. Further, kcat/Km varied less between arginine-containing substrates than for previously reported cathepsin Bs, indicating different substrate specificity of the osteoclast enzyme. Polyclonal antibody raised to a 25 kDa fragment of the enzyme recognized a single 31 kDa band in SDS/PAGE of osteoclast lysates blotted to poly(vinylidene difluoride), adsorbed collagenolytic activity of osteoclast lysates, and stained avian osteoclasts in tissue sections. Degenerate sense- and antisense-oligonucleotide primers, predicted from segments of primary amino acid sequence, amplified a 486 bp DNA fragment; this was cloned and sequenced. Of 162 amino acids encoded, 77% are identical with those of human cathepsin B; hybridization identified a 2.4 kb RNA in osteoclast lysates. We conclude that the major avian osteoclast collagenolytic enzyme is a cathepsin B, whose activity varies from other enzymes of its class.

scholarly journals Actinidain-hydrolyzed Type I Collagen Reveals a Crucial Amino Acid Sequence in Fibril Formation

2010 ◽  
Vol 285 (23) ◽  
pp. 17465-17470 ◽  
Author(s):  
Saori Kunii ◽  
Koichi Morimoto ◽  
Kouhei Nagai ◽  
Takuya Saito ◽  
Kenji Sato ◽  
...  
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Type I Collagen ◽  
Fibril Formation ◽  
Type I

scholarly journals Isolation and Characterization of a Type I Gene Encoding a Light-harvesting Chlorophyll a/b-binding Protein of Photosystem II in Peach

1998 ◽  
Vol 123 (4) ◽  
pp. 493-499 ◽  
Author(s):  
Kyu H. Chung ◽  
Dennis E. Buetow ◽  
Schuyler S. Korban
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Photosystem Ii ◽  
Amino Acid Sequence ◽  
Woody Plants ◽  
Light Harvesting ◽  
Binding Protein ◽  
Transit Peptide ◽  
Type I ◽  
Polyadenylation Sites

A nuclear gene, Lhcb1*Pp1, encoding a light-harvesting chlorophyll a/b-binding protein of photosystem II has been isolated from peach [Prunus persica (L.) Batsch. `Stark Earliglo'] leaf genomic DNA, cloned, and sequenced. This gene encodes a precursor polypeptide of 267 amino acids with a transit peptide of 34 and a type I mature protein of 233 amino acids. The amino acid sequence of the mature polypeptide is 89% to 94% and 80% to 94% similar to those encoded by type I Lhcb genes of annual and other woody plants, respectively. In contrast, the amino acid sequence of the peach transit peptide is less conserved being 47% to 69% similar to those of annual plants and only 17% to 22% similar to those of other woody plants. The peach gene was used as a probe for Lhcb gene expression. Lhcb mRNA is detected in leaves of field-grown trees during June to October. Lhcb mRNA is detected at a high level in leaves of peach shoots grown in tissue culture in the light, but only at a trace level in leaves grown in the dark. Some Lhcb genes appear to be light-modulated in stems. Lhcb1*Ppl contains four potential polyadenylation sites. S1 nuclease analysis detected transcripts of the sizes expected from each of the four polyadenylation sites. All four are found in leaves of light-grown shoots and of field-grown trees throughout the growing season. In contrast, only three are detected in stems of light-grown shoots.

Different responses of biochemical markers of bone resorption to bisphosphonate therapy in Paget disease

1995 ◽  
Vol 41 (11) ◽  
pp. 1592-1598 ◽  
Author(s):  
A Blumsohn ◽  
K E Naylor ◽  
A M Assiri ◽  
R Eastell
Keyword(s):  
Bone Resorption ◽  
Biochemical Markers ◽  
Type I Collagen ◽  
Bisphosphonate Therapy ◽  
Response To Therapy ◽  
Tartrate Resistant Acid Phosphatase ◽  
Type I ◽  
Paget Disease ◽  
Markers Of Bone Resorption ◽  
Total Alkaline

Abstract We examined the response of different biochemical markers of bone resorption to bisphosphonate therapy (400 mg of etidronate daily for 6 months) in mild Paget disease (n = 14). Urinary markers included hydroxyproline (OHP), total (T) and free (F) pyridinolines (Pyds) determined by HPLC, immunoreactive FPyds, immunoreactive TPyds, and the N- and C-terminal telopeptides of type I collage (NTx, CL). Serum measurements included tartrate-resistant acid phosphatase (TRAcP) and the C-terminal telopeptide of type I collagen (ICTP). ICTP and TRAcP showed a minimal response to therapy (% change at 6 months, -13.1 +/- 6.8 and -6.7 +/- 3.4, respectively). The response was greatest for urinary telopeptides (NTx and CL; % change -75.7 +/- 7.5 and -73.4 +/- 8.9, respectively). The response was somewhat greater for TPyds than for FPyds. We conclude that: (a) ICTP and TRAcP are unreliable indicators of changes in bone turnover; (b) oligopeptide-bound Pyds and telopeptide fragments of type I collagen in urine show a somewhat greater response to therapy than do FPyds and may be more sensitive indicators of bone resorption; and (c) as yet no evidence suggests that these markers are substantially better predictors of the clinical response to therapy than serum total alkaline phosphatase or urinary OHP. There are several problems with the interpretation of these measurements in Paget disease, and the clinical utility of these measurements remains uncertain.

Carboxyterminal telopeptide of type I collagen, ICTP, as a marker of matrix degradation in neonatal mouse calvarial bones, in vitro

1992 ◽  
Vol 12 (5) ◽  
pp. 407-411 ◽  
Author(s):  
Östen Ljunggren ◽  
Sverker Ljunghall
Keyword(s):  
Parathyroid Hormone ◽  
Bone Resorption ◽  
Type I Collagen ◽  
Bone Matrix ◽  
Neonatal Mouse ◽  
Type I ◽  
Matrix Degradation ◽  
Dose Responses ◽  
Carboxyterminal Telopeptide

Bone resorption, in vitro, is often measured as the release of prelabelled45Ca from neonatal mouse calvarial bones, or from fetal rat long bones. In this report we describe a technique to measure the breakdown of bone-matrix, in vitro. We also describe a new way to dissect neonatal mouse calvarial bones, in order to obtain large amounts of bone samples. Twelve bone fragments were dissected out from each mouse calvaria and were thereafter cultured in CMRL 1066 culture medium in serum-free conditions in 0.5 cm2 multiwell culture dishes. Matrix degradation after treatment with parathyroid hormone was assessed by measuring the amount of carboxyterminal telopeptide of type I collagen (ICTP) by RIA. The data on matrix degradation was compared to the release of prelabelled45Ca from neonatal mouse calvarial bones. We found that the dose-responses for parathyroid hormone-induced release of prelabelled45Ca and ICTP were identical. In conclusion: RIA-analysis of the ICTP-release is an easy and accurate method to measure degradation of bone-matrix, in vitro. Furthermore, the new dissection technique, described in this report, makes it easy to obtain large amounts of bone samples and thus to perform extensive experiments, e.g. dose-responses for agents that enhance bone resorption.

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