Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.2.1.108 (lactase)
 2,133 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We determined along the small intestine of young and adult rabbits the activities of lactase (LPH) and sucrase (SI), the levels of their cognate mRNAs, and examined the in vitro biosynthesis of LPH and pro-SI. Lactase activity is low in the proximal 1/3 of the intestine, whereas the mRNA levels are high. However, the rates of biosynthesis of the LPH forms correlated well with the steady-state levels of LPH mRNA in all segments, indicating that factor(s) acting post-translationally produce a decline in brush border LPH in the proximal small intestine. These factor(s) are not involved in the processing of pro-LPH to mature LPH, since the relative amounts of the various forms of LPH are almost the same along the small intestine. Unexpectedly, we find that also for SI the ratio of activity to mRNA is low in proximal intestine. The biosynthesis of pro-SI correlates with the steady-state levels of its mRNA. Hence, the steady-state levels of LPH and SI along the small intestine are regulated both by mRNA levels and by posttranslational factor(s).
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PMID:The levels of lactase and of sucrase-isomaltase along the rabbit small intestine are regulated both at the mRNA level and post-translationally. 144 47

Very sensitive procedures were developed for the parallel determination of intestinal lactase (LPH) activity and the cognate mRNA. Between 14 and 20 weeks of gestation, lactase activity is low and varies only slightly; at 37 weeks, a relatively high level of activity is observed. The amounts of LPH mRNA correlates with the enzymatic activity (r = 0.64). Culture of fetal jejunal explants for 5 days induces by itself a 2-fold increase in LPH mRNA, without any significant change in lactase enzymatic activity. This increase may reflect the loss of a negative transcriptional regulation operative in vivo, and suggests an additional post-transcriptional regulatory component. The addition of hydrocortisone (50 ng/ml) during culture induces a doubling of lactase activity without variation in LHP mRNA, indicating a post-transcriptional modulation by hydrocortisone. The intestinal lysosomal acid beta-galactosidase activity was shown to be unaffected by hydrocortisone treatment. This observation clearly illustrates that the two intestinal beta-galactosidases are regulated differently. Our results suggests a complex developmental regulation of human intestinal lactase and that the perinatal increase in lactase activity could be modulated at a post-transcriptional level by hydrocortisone.
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PMID:The expression of lactase enzymatic activity and mRNA in human fetal jejunum. Effect of organ culture and of treatment with hydrocortisone. 156 81

We report the primary structures of human and rabbit brush border membrane beta-glycosidase complexes (pre-pro-lactase-phlorizin hydrolase, or pre-pro-LPH, EC 3.2.1.23-62), as deduced from cDNA sequences. The human and rabbit primary translation products contain 1927 and 1926 amino acids respectively. Based on the data, as well as on peptide sequences and further biochemical data, we conclude that the proteins comprise five domains: (i) a cleaved signal sequence of 19 amino acids; (ii) a large 'pro' portion of 847 amino acids (rabbit), none of which appears in mature, membrane-bound LPH; (iii) the mature LPH, which contains both the lactase and phlorizin hydrolase activities in a single polypeptide chain; (iv) a membrane-spanning hydrophobic segment near the carboxy terminus, which serves as membrane anchor; and (v) a short hydrophilic segment at the carboxy terminus, which must be cytosolic (i.e. the protein has an Nout-Cin orientation). The genes have a 4-fold internal homology, suggesting that they evolved by two cycles of partial gene duplication. This repetition also implies that parts of the 'pro' portion are very similar to parts of mature LPH, and hence that the 'pro' portion may be a water-soluble beta-glycosidase with another cellular location than LPH. Our results have implications for the decline of LPH after weaning and for human adult-type alactasia, and for the evolutionary history of LPH.
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PMID:Complete primary structure of human and rabbit lactase-phlorizin hydrolase: implications for biosynthesis, membrane anchoring and evolution of the enzyme. 246 Mar 43

The biosynthesis and maturation of the human intestinal lactase-phlorizin hydrolase (LPH; EC 3.2.1.23-3.2.1.62) has been studied in cultured intestinal biopsies and mucosal explants. Short time pulse labelling revealed on high mannose intermediate of Mr 215,000 which was converted upon endo-beta-N-acetylglucosaminidase H (endo-H) digestion to a polypeptide of Mr 200,000. The brush border form of LPH was revealed after longer pulse periods and has Mr 160,000. It possesses mainly complex oligosaccharide chains and, owing to its partial endo-H sensitivity, at least one chain of the high mannose type. Leupeptin partially inhibited the appearance of the Mr-160,000 polypeptide. Monensin treatment of biopsies resulted in the modification of the Mr-160,000 species to the Mr-140,000 molecule, which was endo-H sensitive. Pulse-chase analysis indicated a slow post-translational processing of the high mannose precursor (Mr 215,000) to yield the mature brush-border form (Mr 160,000) of LPH. Our results further indicate that LPH is synthesized as a single polypeptide precursor which is intracellularly cleaved to yield the mature brush border of LPH. The data presented suggest that this cleavage occurs during the translocation of the molecule across the Golgi complex.
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PMID:Biosynthesis and maturation of lactase-phlorizin hydrolase in the human small intestinal epithelial cells. 310 75

Enterocytes are the major epithelial cell type of the small intestine. Their capacity to secret, absorb and digest specific ions and nutrients is dependent on their position along the length of the small intestine as well as their stage of development as they migrate and differentiate along the crypt-villus axis. In order to further understand the molecular processes that regulate enterocyte differentiation and function, this study has compared the levels of six mRNA species produced by genes expressed in rabbit enterocytes; specifically, the multidrug resistance (MDR1) gene encoding the 170-kDa P-glycoprotein, CaBP 9k, which encodes a putative intracellular calcium buffer, calbindin, LPH, APN, and AP which encode the brush-border hydrolases lactase-phlorizin hydrolase, aminopeptidase N and alkaline phosphatase, respectively, and SGLT1, encoding the brush border Na(+)-glucose cotransporter. The level of each mRNA species has been mapped along the small intestine using quantitative in situ hybridisation. This has revealed characteristic regional variations in the abundance of each of the mRNAs, supporting the opinion that there is a strong genetic component to the maintenance of gradients in epithelial function along the length of the small intestine. Analysis of the cellular accumulation of mRNA during enterocyte migration along the crypt-villus axis, over gut-associated lymphoid tissue, and at epithelial boundaries, has, by contrast, established a clear correlation in the expression of these genes. These data illustrate the dynamics of enterocyte gene expression, thereby providing an insight into the molecular mechanisms which co-ordinate the events of cell transformation that underlie functional differences between the epithelial populations of the small intestine.
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PMID:Parallel patterns of cell-specific gene expression during enterocyte differentiation and maturation in the small intestine of the rabbit. 758 2

The pro-region of intestinal lactase-phlorizin hydrolase (LPH alpha) has been proposed to be important for the correct folding of pro-LPH and mature LPH (LPH beta). In this communication, analysis of the catalytic function of the LPH alpha pro-region is presented. Expression of a cDNA encoding LPH alpha in COS-1 cells reveals a polypeptide that does not hydrolyse lactose. Likewise, no lactase activity is detected in LPH alpha purified from trypsin-treated pro-LPH. Mixing of LPH alpha and LPH beta does not lead to the activation of the latter. We conclude that LPH alpha does not contribute to the lactase activity despite the strong homologies with mature LPH beta. LPH alpha may play an important role as an intra-molecular chaperone.
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PMID:The pro-region of human intestinal lactase-phlorizin hydrolase is enzymatically inactive towards lactose. 762 35

Milk lactose is hydrolysed to D-galactose and D-glucose in the small intestine of mammals by the lactase-phlorizin hydrolase complex (LPH, EC 3.2.1.23-62). Lactase activity has broad substrate selectivity and several glycosides are substrates. Recently, using the monodeoxy derivatives of methyl beta-lactoside (1), we have shown the importance of each hydroxyl group in the substrate molecule concerning the interaction with the enzyme. Now we have studied the corresponding O-methyl derivatives, as well as some of the halo derivatives of 1. We have found that the enzyme presents steric restrictions to the recognition of substrates modified in the galactose moiety. In contrast, the binding site for the aglycon part of the substrate is looser. On the other hand, we have previously shown that HO-3' and HO-6 were important for the recognition of the substrate by the enzyme. Now we have found that the corresponding fluorine derivatives are not, or very poorly, recognized. This suggests that the HO-3' and HO-6 participate, as donors, in hydrogen bonds in the interaction with the enzyme.
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PMID:Substrate specificity of small-intestinal lactase: study of the steric effects and hydrogen bonds involved in enzyme-substrate interaction. 764 81

Various cDNAs coding for part or all of human pre-pro lactase-phlorizin hydrolase (pre-proLPH) were transfected into COS cells and the subcellular location of the lactase-related proteins assessed. Only the complete proLPH reached the plasma membrane. LPH without the pro sequence, and a construct containing the pro sequence and the lactase domain of mature LPH, accumulated intracellularly; the pro sequence with no mature domain was secreted. We conclude that the pro sequence is important for LPH to be transported to the cell surface.
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PMID:The pro sequence of lactase-phlorizin hydrolase is required for the enzyme to reach the plasma membrane. An intramolecular chaperone? 822 50

The promoter activity of the upstream region of the rat small intestinal lactase-phlorizin hydrolase gene has been analysed by transfection in the human colon cancer cell line Caco-2. A 0.9 kb mRNA, corresponding to the CAT reporter gene, was synthesized from the transcription start site of the LPH gene. The rate of expression, determined by semi-quantitative RT-PCR, was very low, and depended on the length of the promoter fragment in front of the reporter gene. By immunocytology, we found that the low level of expression resulted from the low number of cells (about 1%) in which CAT was produced. The endogenous lactase was present in 10-20% of the cells in culture, and evidence is provided that most cells that expressed CAT did not co-express the endogenous lactase. We conclude from this study that the rat small intestinal LPH gene promoter is active in the human Caco-2 colon cancer cells. Hence Caco-2 cells constitute an in vitro model to analyse the basic molecular mechanisms involved in the gene transcription of intestinal digestive enzymes. Yet, the mosaic expression of the endogenous lactase and of the reporter gene under the control of the rat LPH gene promoter, suggests that Caco-2 cells may present specific regulatory mechanisms of expression of small intestinal enzymes, possibly in relation to their tumourous origin.
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PMID:Activity of the rat lactase gene promoter in transfected human colon cancer cells. 857 90

Human lactase-phlorizin hydrolase (LPH, EC 3.2.1.23/62) is synthesized as a single-chain precursor glycoprotein (pro-LPH) with a relative molecular mass of just over 200 kDa. Maturation to the mature enzyme (m-LPH, 160 kDa) occurs after passage of pro-LPH through the Golgi complex and involves the proteolytic removal of a 849 amino acid propeptide. The role of this propeptide as well as its removal is not fully understood and the proteolytic enzyme or enzymes involved are unknown. We studied the potential role of five different members of the family of subtilisin-like proprotein processing proteases in the maturation process of human LPH using a vaccinia virus based coexpression system in pig kidney PK(15) cells. Infected/transfected PK(15) cells expressed full-length pro-LPH but no maturation to m-LPH was observed. Coexpression of human pro-LPH with human furin, human PC1/PC3, human PC2, human PACE4 and mouse PC6A in PK(15) cells did not result in maturation of the enzyme. Cleavage and secretion of von Willebrand factor precursor (pro-vWF) was used as a positive control. None of the five proprotein processing proteases tested were capable of cleaving human pro-LPH, strongly suggesting that they are not involved in the maturation of this enzyme.
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PMID:Human lactase-phlorizin hydrolase is not processed by furin, PC1/PC3, PC2, PACE4 and PC5/PC6A of the family of subtilisin-like proprotein processing proteases. 866 47


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