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D J Prockop

Publications and source records attributed to D J Prockop.

At least 91 records · Page 5Linked to original sources
PubMed1996

Molecular cloning, expression and chromosomal localization of a human gene encoding a 33 kDa putative metallopeptidase (PRSM1).

The zincins are a superfamily of structurally-related Zn(2+)-binding metallopeptidases which play a major role in a wide range of biological processes including pattern formation, growth factor activation and extracellular matrix synthesis and degradation. In this paper we report the identification and complete primary structure of a novel 33 kDa protein which contains the zinc-binding HEXXH motif found in the zincin superfamily. We have named this novel protein PRSM1 (PRoteaSe, Metallo, number 1). The gene was identified by the immunoscreening of a human placental cDNA library using polyclonal antibodies raised to the 70 kDa human matrix metalloendopeptidase, type III procollagen N-proteinase [Halila, R. and Peltonen, L. (1986) Purification of human procollagen type III N-proteinase from placenta and preparation of antiserum. Biochem. J. 239, 47-52]. The protein is found in placenta and cultured osteosarcoma cells. PRSM1 could share sequence homology with the type III procollagen N-proteinase. The prsm1 gene is represented once in the human genome and is localized on chromosome 16 (q24.3).

Amino Acid Sequence↗
PubMed1996

Assembly in vitro of thin and thick fibrils of collagen II from recombinant procollagen II. The monomers in the tips of thick fibrils have the opposite orientation from monomers in the growing tips of collagen I fibrils.

Human type II procollagen was prepared in a recombinant system and cleaved to pC-collagen II by procollagen N-proteinase. The pC-collagen II was then used as a substrate to generate collagen II fibrils by cleavage with procollagen C-proteinase at 37 degrees C. Electron microscopy of the fibrils demonstrated that, at the early stages of fibril assembly, very thin fibrils were formed. As the system approached equilibrium over 7-12 h, however, the thin fibrils were largely but not completely replaced by thick fibrils that had diameters of about 240 nm and a distinct D-period banding pattern. One typical fibril was photographed and analyzed in its entirety. The fibril was 776 D-periods (52 microM) long. It had a central shaft with a uniform diameter that was about 516 D-periods long and two tips of about 100 D-periods each. Most of the central shaft had a symmetrical banding pattern flanked by two transition regions of about 30 D-periods each. Measurements by scanning transmission electron microscopy demonstrated that the mass per unit length from the tips to the shafts increased linearly over approximately 100 D-periods from the fibril end. The linear increase in mass per unit length was consistent with previous observations for collagen I fibrils and established that the tips of collagen II also had a near paraboloidal shape. However, the orientation of monomers in the tips differed from the tips of collagen I fibrils in that the C termini instead of the N termini were directed toward the tips. The thin fibrils that were present at early stages of assembly and at equilibrium were comparable to the collagen II fibrils seen in embryonic tissues and probably represented intermediates on the pathway of thick fibrils formation. The results indicated that the molecular events in the self-assembly of collagen II fibrils are apparently similar to those in self-assembly of collagen I fibrils, but that there are also important differences in the structural information contained in collagen I and collagen II monomers.

Cell Line↗
PubMed1996

A-2-->G transition at the 3' acceptor splice site of IVS17 characterizes the COL2A1 gene mutation in the original Stickler syndrome kindred.

Hereditary progressive arthro-ophthalmopathy, or "Stickler syndrome," is an autosomal dominant osteochondrodysplasia characterized by a variety of ocular and skeletal anomalies which frequently lead to retinal detachment and precocious osteoarthritis. A variety of mutations in the COL2A1 gene have been identified in "Stickler" families; in most cases studied thus far, the consequence of mutation is the premature generation of a stop codon. We report here the characterization of a COL2A1 gene mutation in the original kindred described by Stickler et al. [1965]. Conformational sensitive gel electrophoresis (CSGE) [Ganguly et al., 1993] was used to screen for mutations in the entire COL2A1 gene in an affected member from the kindred. A prominent heteroduplex species was noted in the polymerase chain reaction (PCR) product from a region of the gene including exons 17 to 20. Direct sequencing of PCR-amplified genomic DNA resulted in the identification of a base substitution at the A-2 position of the 3' splice acceptor site of IVS17. Sequencing of DNA from affected and unaffected family members confirmed that the mutation segregated with the disease phenotype. Reverse transcriptase-PCR analysis of poly A+ RNA demonstrated that the mutant allele utilized a cryptic splice site in exon 18 of the gene, eliminating 16 bp at the start of exon 18. This frameshift eventually results in a premature termination codon. These findings are the first report of a splice site mutation in classical Stickler syndrome and they provide a satisfying historical context in which to view COL2A1 mutations in this dysplasia.

Adolescent↗
PubMed1996

The C-proteinase that processes procollagens to fibrillar collagens is identical to the protein previously identified as bone morphogenic protein-1.

Bone morphogenic protein-1 (BMP-1) was originally identified as one of several BMPs that induced new bone formation when implanted into ectopic sites in rodents. BMP-1, however, differed from other BMPs in that it its structure was not similar to transforming growth factor beta. Instead, it had a large domain homologous to a metalloendopeptidase isolated from crayfish, an epidermal growth-factor-like domain, and three regions of internal sequence homology referred to as CUB domains. Therefore, BMP-1 was a member of the "astacin families" of zinc-requiring endopeptidases. Many astacins have been shown to play critical roles in embryonic hatching, dorsal/ventral patterning, and early developmental decisions. Here, we have obtained amino acid sequences and isolated cDNA clones for procollagen C-proteinase (EC 3.4.24.19), an enzyme that is essential for the processing of procollagens to fibrillar collagens. The results demonstrate that procollagen C-proteinase is identical to BMP-1.

Amino Acid Sequence↗
PubMed1996

Substitution of arginine for glycine at position 154 of the alpha 1 chain of type I collagen in a variant of osteogenesis imperfecta: comparison to previous cases with the same mutation.

A substitution of arginine for glycine at amino acid position 154 of the alpha 1(I) collagen chain was found in a father and his three children. The phenotype of the patients includes manifestations of types I and III/IV osteogenesis imperfecta, but appears to be milder than that of the previously described two unrelated patients that had the identical mutation in the alpha 1(I) collagen chain. The variability in the phenotype raises the possibility of epistatic loci or environmental effects on expression of the disorder.

Adult↗
PubMed1996

Perspectives on the synthesis and application of triple-helical, collagen-model peptides.

Collagens can be distinguished from other proteins based on their triple-helical structure. Synthetic peptide models have been developed to better understand the triple helix structurally and to evaluate the triple helix as a recognition element for biological processes. Associated triple-helical peptides were first designed and assembled by solid-phase methodology in the late 1960s. Such peptides were used for triple-helical structural characterization by CD, nmr, and ir spectroscopies, and x-ray crystallography, and for studying the structural preferences of hydroxylases. In the late 1970s, methods were developed for covalently linking the three strands of triple-helical peptides. One benefit of "branched" peptides was the enhancement of triple-helical thermal stability. The incorporation of specific collagen sequences into thermally stable synthetic triple helices in the early 1990s has allowed for the mechanistic investigation of collagen-mediated cell adhesion and platelet aggregation. In time, discriminatory therapeutics may result from the continued exploration and further understanding of the biological effects of collagen primary, secondary, and tertiary structures via triple-helical peptide models.

Amino Acid Sequence↗
PubMed1996

Direct sequencing of PCR products derived from cDNAs for the pro alpha 1 and pro alpha 2 chains of type I procollagen as a screening method to detect mutations in patients with osteogenesis imperfecta.

More than 150 mutations in the genes for type I procollagen have been found in unrelated patients with osteogenesis imperfecta (OI), but mutations have been difficult to define in many patients with the mildest forms of the disease. Here, we have used robotically automated sequencing of the cDNAs for type I procollagen to screen for mutations in 12 patients suspected of having nonlethal OI (types I, III, and IV). Single base mutations that changed codons for obligate glycine residues were found in seven of the patients. Altogether, we analyzed 4,379 bp of sequences of both alleles of the pro alpha 1 (I) collagen (8,758 bp of allelic sequences) and 4,200 bp of sequences of both alleles of the pro alpha 2(I) collagen (8,400 bp of allelic) from each patient.

Adult↗
PubMed1996

Vitamin D receptor genotype is not associated with bone mineral density in three ethnic/regional groups.

We report a cross-sectional study of 48 men, 56 premenopausal women, and 80 postmenopausal women who were of three ethnic/regional backgrounds: southern European (Greek, Italian), eastern European (Jewish, Polish, Hungarian), and western European (French, British). We determined bone mineral density (BMD) at four skeletal sites and assessed the vitamin D receptor (VDR) genotype by the Bsml restriction site polymorphism. Age and body mass index had significant effects on BMD by multiple regression analysis. In addition, ethnic/regional group had a significant effect on spinal BMD in premenopausal females (P = 0.014) and in males (P = 0.039). However, VDR genotype had no significant effect on BMD in any of the three study groups.

Bone Density↗
PubMed1996

The mouse col11a2 gene. Some transcripts from the adjacent rxr-beta gene extend into the col11a2 gene.

Type XI collagen is present in small amounts in cartilage, together with small amounts of type IX and type V collagens and large amounts of type II collagen. Here, primers based on the nucleotide sequences of partial human cDNAs and mouse genomic DNAs that were analyzed by other investigators were used to isolate a cDNA for the mouse col11a2 gene. Cosmid clones for the mouse col11a2 gene were isolated, and 12.4 kb of the nucleotide sequences were defined. Analysis of the genomic sequences identified three exons in the mouse gene that were recently shown to undergo alternative splicing (Tsumaki and Kimura, J. Biol. Chem. 270, 2372-2378, 1995; Zhidkova et al., J. Biol. Chem. 270, 94886-9493, 1995). In addition, analysis of the cosmid clones revealed that the 5' end of the mouse col11a2 gene was located head-to-tail with the mouse retinoic X receptor beta gene. RT-PCR assays demonstrated that some transcripts from the retinoic X receptor beta gene extend into the col11a2 gene. Therefore, there may be coordinate expression of the two genes.

Animals↗
PubMed1996

Mineral changes in a transgenic mouse model for osteogenesis imperfecta.

A line of transgenic mice has been investigated that expressed moderate levels of an internally deleted human gene for the pro alpha 1(I) chain of type I procollagen to determine if they would make a good model for osteogenesis imperfecta (brittle bone disease). Previous workers have reported extensive fracturing in these mice, with femurs that were shorter and bone that had decreased ash weight, mineral and collagen content. These workers demonstrated increased brittleness in the bone by biomechanical measurements. The molar calcium to phosphorus ratio in bone from patients with osteogenesis imperfecta has previously been reported to be lower than that in normal human bone. Mineral changes were observed at the ultrastructural level in these mice and were comparable with those seen in patients with osteogenesis imperfecta. Bone from both transgenic and normal littermate mice was examined to determine if any similarity with the data for human osteogenesis imperfecta could be drawn. X-ray microanalysis of bone mineral demonstrated a lower calcium to phosphorus molar ratio in transgenic mouse bone than in normal littermates. Fourier-transform infra-red spectroscopy confirmed that the mineral present was apatitic in nature despite the lower calcium to phosphorus molar ratio. Multiple fracture calluses were present on the ribs and on the long bones of the transgenic mice; this was absent in normal littermates. This mouse model may lead to a better understanding of the underlying pathology resulting in fragile bones in osteogenesis imperfecta.

Animals↗
PubMed1996

Osteopenia in 37 members of seven families: analysis based on a model of dominant inheritance.

BACKGROUND: The genetic factors involved in determining bone mineral density (BMD) have not been fully elucidated. We have begun genetic linkage analysis of seven families in which many members are osteopenic, in order to identify chromosomal loci that are potentially involved in determining BMD. MATERIALS AND METHODS: Spine BMD was measured in 143 members of seven kindred with familial osteopenia. The absolute BMD values for the spine (L2-L4) were converted to the age-, gender-, and weight-adjusted Z scores, and this corrected value was used as the quantitative trait on which to base subsequent genetic analyses. Simulations of linkage were performed in order to determine the information content of the pedigree set, and actual linkage analysis was conducted using polymorphic markers either within or near three candidate loci: COL1A1, COL1A2, and vitamin D receptor (VDR). RESULTS: The distribution of the corrected Z scores was bimodal (p = 0.001) suggesting a monogenic mode of inheritance of the low BMD trait. Simulation of linkage analysis suggested that the family data set was sufficient to detect linkage under a single major gene model. Actual linkage analysis did not support linkage to the three candidate loci. In addition, the VDR genotype was not statistically associated with low bone density at the spine. CONCLUSIONS: Loci other than COL1A1, COL1A2 and VDR are very likely responsible for the low BMD trait observed in these families. These families are suitable for a genome-wide screen using microsatellite repeats in order to identify the loci that are involved in osteopenia.

Analysis of Variance↗