The third activity for lysyl hydroxylase 3: galactosylation of hydroxylysyl residues in collagens in vitro

Wang, Chunguang; Luosujärvi, Hanne; Heikkinen, Jari; Risteli, Maija; Uitto, Lahja; Myllylä, Raili

doi:10.1016/s0945-053x(02)00071-9

Cited by 94 publications

(123 citation statements)

References 38 publications

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“…In addition to LH activity, LH3, but not LH1 or LH2, also possesses collagen glucosyltransferase activity and very small amounts of collagen galactosyltransferase activity (11)(12)(13), implying that this protein may be able to perform all of the reactions involved in the formation of hydroxylysine and its carbohydrate units. The level of collagen galactosyltransferase activity was not decreased in the LH3 null embryos, but collagen glucosyltransferase activity had decreased to Ϸ15% in the null embryos, indicating that this activity of LH3 is of real biological significance and may contribute to the phenotype in null mice.…”

Section: Discussionmentioning

confidence: 99%

“…This protein differs from the other two lysyl hydroxylase isoenzymes in that it also possesses relatively low levels of collagen glucosyltransferase activity and very small amounts of collagen galactosyltransferase activity in addition to the lysyl hydroxylase activity (11)(12)(13). In this study, we generated mice with targeted inactivation of the LH3 (Plod3) gene and show that LH3 is essential for the synthesis of type IV collagen during early development and, hence, for the stability of basement membranes (BMs).…”

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confidence: 99%

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Premature aggregation of type IV collagen and early lethality in lysyl hydroxylase 3 null mice

Rautavuoma

Takaluoma

Sormunen

et al. 2004

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

102

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Collagens carry hydroxylysine residues that act as attachment sites for carbohydrate units and are important for the stability of crosslinks but have been regarded as nonessential for vertebrate survival. We generated mice with targeted inactivation of the gene for one of the three lysyl hydroxylase isoenzymes, LH3. The null embryos developed seemingly normally until embryonic day 8.5, but development was then retarded, with death around embryonic day 9.5. Electron microscopy (EM) revealed fragmentation of basement membranes (BMs), and immuno-EM detected type IV collagen within the dilated endoplasmic reticulum and in extracellular aggregates, but the typical BM staining was absent. Amorphous intracellular and extracellular particles were also seen by collagen IV immunofluorescence. SDS͞PAGE analysis demonstrated increased mobilities of the type IV collagen chains, consistent with the absence of hydroxylysine residues and carbohydrates linked to them. These results demonstrate that LH3 is indispensable for biosynthesis of type IV collagen and for BM stability during early development and that loss of LH3's functions leads to embryonic lethality. We propose that the premature aggregation of collagen IV is due to the absence of the hydroxylysine-linked carbohydrates, which thus play an essential role in its supramolecular assembly. L ysyl hydroxylase (LH, EC 1.14.11.4, Plod) catalyzes the hydroxylation of lysine residues in -X-Lys-Gly-triplets in collagens and other proteins with collagen-like sequences (1). The hydroxylysine residues formed have two important functions: (i) they serve as attachment sites for carbohydrates, either monosaccharide galactose or disaccharide glucosylgalactose, and (ii) they have an important role in the stabilization of intermolecular collagen crosslinks that provide tensile strength and mechanical stability for the collagen fibrils. The extent of lysine hydroxylation and hydroxylysine glycosylation varies between collagen types. Nearly 90% of all lysines are hydroxylated and hydroxylysines glycosylated in types IV and VI collagen, whereas Ͻ20% of the lysines are hydroxylated in type III collagen (2). The functions of the hydroxylysine-linked carbohydrates are poorly understood, but in the case of fibril-forming collagens, they are thought to regulate the formation and morphology of the fibrils (1, 3).Three human LH isoenzymes have been identified (1, 4-6). Mutations in the LH1 gene lead to the kyphoscoliotic type of Ehlers-Danlos syndrome, which is characterized by scoliosis, joint laxity, skin fragility, ocular manifestations, and severe muscle hypotonia (7-9). Mutations in the LH2 gene have recently been reported in two families with Bruck syndrome, which is characterized by fragile bones, joint contractures, scoliosis, and osteoporosis (10).No mutations have been characterized in the gene for the isoenzyme LH3, and its in vivo functions have not yet been elucidated. This protein differs from the other two lysyl hydroxylase isoenzymes in that it also possesses relatively low levels...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Premature aggregation of type IV collagen and early lethality in lysyl hydroxylase 3 null mice

Rautavuoma

Takaluoma

Sormunen

et al. 2004

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

102

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“…9 LH3 differs from LH1 and LH2 in that it is multifunctional and able to catalyze, in addition to Lys hydroxylation, sugar transfer reactions, the subsequent steps in the formation of glucosylgalactosyl-Hyl residues. [12][13][14] Only one isoform for LH is present in lower species such as Caenorhabditis elegans. 9,15 This ancestral C. elegans LH is also able to glycosylate Hyl residues and, thus, is functionally similar to LH3.…”

Section: Introductionmentioning

confidence: 99%

“…9,15 This ancestral C. elegans LH is also able to glycosylate Hyl residues and, thus, is functionally similar to LH3. 9,13 Recently, Salo et al 16 found that LH3 is located in two compartments in tissues, in the endoplasmic reticulum and the extracellular space, and the partitioning varies with tissue type. Thus, LH3 may be functional both inside and outside of the cell.…”

Section: Introductionmentioning

confidence: 99%

Development of a convenient peptide‐based assay for lysyl hydroxylase

Čudić¹,

Patel²,

Lauer‐Fields³

et al. 2008

Biopolymers

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Hydroxylysine (Hyl) is a posttranslationally modified amino acid found mainly in collagens, the most abundant protein in mammals. Lysyl hydroxylase (LH) catalyzes the hydroxylation of the C‐5 position of a Lys residue, resulting in the production of Hyl. Mechanistically, LH incorporates one oxygen atom into both Lys and 2‐oxoglutarate; the latter is decarboxylated to form succinate and CO2. To develop a convenient, RP‐HPLC based LH assay, we used Fmoc solid‐phase methodology to synthesize three different peptides designed as LH substrates and one peptide corresponding to an LH product. Peptides were characterized by RP‐HPLC, MALDI‐TOF mass spectrometry and CD spectroscopy. Separation of peptides was examined under a variety of RP‐HPLC conditions. The best results were achieved using peptide derivatization (1‐anthroylnitrile for organic phase and dansyl chloride for aqueous phase) prior to RP‐HPLC analysis. The products (di‐ and tetra‐substituted Lys‐ and Hyl‐containing peptides) were well resolved by RP‐HPLC. The resolution of each peak allows for quantification of peak areas, which in turn, when examined as a function of time, can be utilized for studying the kinetics of LH catalyzed reactions. Most significantly, the RP‐HPLC assay directly monitors the Hyl containing product. Prior LH assay methods are multi‐step, require radio‐labeled substrates, and/or measure depletion of 2‐oxoglutarate or formation of CO2. Since the LH reaction with 2‐oxoglutarate is uncoupled from Lys hydroxylation, the most accurate assay of LH activity should monitor the formation of Hyl. © 2007 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 90: 330–338, 2008.This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com

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“…Recently, mutations in human lh2 (plod2) were linked to Bruck syndrome, distinguished by fragile bones and contractures of the large joints (Ha-Vinh et al, 2004;van der Slot et al, 2003). LH3, in addition to exhibiting lysyl hydroxylase activity, is the only LH enzyme to possess additional glycosyltransferase activities that serve to further modify hydroxylysine residues to galactosylhydroxylysine and glucosylgalactosylhydroxylysine (Heikkinen et al, 2000;Rautavuoma et al, 2002;Wang et al, 2002). Although mutations in human lh3 (plod3) have not yet been identified, a mouse knockout reveals this gene has an embryonic lethal phenotype and is required for type IV collagen secretion (Rautavuoma et al, 2004;Ruotsalainen et al, 2006).…”

Section: Introductionmentioning

confidence: 99%

Genomic structure and embryonic expression of zebrafish lysyl hydroxylase 1 and lysyl hydroxylase 2

Schneider

Granato

2007

Matrix Biology

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Collagen biosynthesis in both invertebrates and vertebrates is critically dependent upon the activity of lysyl hydroxylase (LH) enzymes. In humans, mutations in the genes encoding LH1 and LH2 have been shown to cause two distinct connective tissue disorders, Ehlers-Danlos (Type VIA) and Bruck syndromes. While the biochemical properties of these enzymes have been intensively studied, their embryonic patterns of expression and developmental roles remain unknown. We now present the cloning and analyses of the genes encoding LH1 and LH2 in the zebrafish, Danio rerio. We find these genes to be similarly organized to other vertebrate lh (plod) genes, including the presence of an alternatively spliced exon in lh2. We also examine the mRNA expression patterns of lh1 and lh2 during embryogenesis and find them to exhibit unique and dynamic patterns of expression. These results strongly suggest that LH enzymes are not merely housekeeping enzymes, but play distinct developmental roles. The identification of these genes in the zebrafish, a genetic model organism whose development is well characterized, now provides the basis for the establishment of the first animal models for both Ehlers-Danlos (Type VIA) and Bruck syndromes.

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The third activity for lysyl hydroxylase 3: galactosylation of hydroxylysyl residues in collagens in vitro

Cited by 94 publications

References 38 publications

Premature aggregation of type IV collagen and early lethality in lysyl hydroxylase 3 null mice

Premature aggregation of type IV collagen and early lethality in lysyl hydroxylase 3 null mice

Development of a convenient peptide‐based assay for lysyl hydroxylase

Genomic structure and embryonic expression of zebrafish lysyl hydroxylase 1 and lysyl hydroxylase 2

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