Betaine-homocysteine S-methyltransferase (human)

Skiba, William E.; Wells, M S; Mangum, John H.; Awad, William M.

doi:10.1016/0076-6879(87)43067-x

Cited by 13 publications

(3 citation statements)

References 15 publications

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“…The K m values obtained for the recombinant wild-type protein differed from those published previously for the rat liver purified BHMT [52,53], being 9-and 3-fold higher for Hcy and betaine respectively. In fact, the K m values obtained for Hcy are closer to those published for the human liver purified enzyme [54] than to those for the recombinant human BHMT [13]. Regarding betaine, the K m values differed markedly from those published for the human liver enzyme, which are approx.…”

Section: Spectra (%)contrasting

confidence: 61%

Active-site-mutagenesis study of rat liver betaine-homocysteine S-methyltransferase

González

Campillo

Garrido

et al. 2003

Biochemical Journal

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A site-directed-mutagenesis study of putative active-site residues in rat liver betaine-homocysteine S-methyltransferase has been carried out. Identification of these amino acids was based on data derived from a structural model of the enzyme. No alterations in the CD spectra or the gel-filtration chromatography elution pattern were observed with the mutants, thus suggesting no modification in the secondary structure content or in the association state of the proteins. All the mutants obtained showed a reduction of the enzyme activity, the most dramatic effect being that of Glu(159), followed by Tyr(77) and Asp(26). Changes in affinity for either of the substrates, homocysteine or betaine, were detected when substitutions were performed of Glu(21), Asp(26), Phe(74) and Cys(186). Interestingly, Asp(26), postulated to be involved in homocysteine binding, has a strong effect on affinity for betaine. The relevance of these results is discussed in the light of very recent structural data obtained for the human enzyme.

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Section: Spectra (%)contrasting

confidence: 61%

Active-site-mutagenesis study of rat liver betaine-homocysteine S-methyltransferase

González

Campillo

Garrido

et al. 2003

Biochemical Journal

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“…5), which strongly suggests that the cleavage of p44 to yield p35 occurs in the cytosol. It has been reported previously (46) that the partial degradation of BHMT to produce low molecular mass forms occurs during purification and that this degradation can be inhibited by dimethylglycine and homocysteine, the product and substrate, respectively, of the BHMT reaction. In accordance with this observation, we performed immunoblotting to confirm that the addition of dimethylglycine and homocysteine (each 2 mM) to the homogenization buffer markedly reduces the p35 level in liver homogenates (data not shown).…”

Section: Fig 2 Golgi/endosome Marker Proteins Rab Gtp-binding Protmentioning

confidence: 99%

Autolysosomal Membrane-associated Betaine Homocysteine Methyltransferase

Ueno¹,

Ishidoh²,

Mineki³

et al. 1999

Journal of Biological Chemistry

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We compared the membrane proteins of autolysosomes isolated from leupeptin-administered rat liver with those of lysosomes. In addition to many polypeptides common to the two membranes, the autolysosomal membranes were found to be more enriched in endoplasmic reticulum lumenal proteins (protein-disulfide isomerase, calreticulin, ER60, BiP) and endosome/Golgi markers (cation-independent mannose 6-phosphate receptor, transferrin receptor, Golgi 58-kDa protein) than lysosomal membranes. The autolysosomal membrane proteins include three polypeptides (44, 35, and 32 kDa) whose amino-terminal sequences have not yet been reported. Combining immunoblotting and reverse transcriptase-polymerase chain reaction analyses, we identified the 44-kDa peptide as the intact subunit of betaine homocysteine methyltransferase and the 35-and 32-kDa peptides as two proteolytic fragments. Pronase digestion of autolysosomes revealed that the 44-kDa and 32-kDa peptides are present in the lumen, whereas the 35-kDa peptide is not. In primary hepatocyte cultures, the starvation-induced accumulation of the 32-kDa peptide occurs in the presence of E64d, showing that the 32-kDa peptide is formed from the sequestered 44-kDa peptide during autophagy. The accumulation is induced by rapamycin but completely inhibited by wortmannin, 3-methyladenine, and bafilomycin. Thus, detection of the 32-kDa peptide by immunoblotting can be used as a streamlined assay for monitoring autophagy.Autophagy is a universal process by which cellular proteins are degraded via a lysosomal/vacuolar system. Depending on the extracellular nutrient conditions, the rate of autophagic protein degradation fluctuates between 1-1.5% and 4 -5% of total cell proteins per hour (1, 2). There are two pathways of autophagy, microautophagy and macroautophagy (for reviews, see Refs. 3 and 4). In microautophagy, relatively small portions of the cytoplasm are directly enclosed by invaginating lysosomal membranes for subsequent sequestration and degradation. Degradation of bulky cell constituents by the lysosome/vacuole system occurs via macroautophagy. In the initial step of macroautophagy, various cytosolic proteins, as well as cytoplasmic organelles such as mitochondria, endoplasmic reticulum (ER), 1 and peroxisomes, are sequestered in the lumen of double-membraned autophagosomes. Autophagosomes then fuse with endosomes or lysosomes to become mature, single-membraned autolysosomes. Acidification of the lumen and acquisition of lysosomal hydrolytic enzymes enable this specialized membrane system to degrade sequestered cytoplasmic components.The origin of the autophagosomal membrane is a subject of controversy. Extensive morphological analyses by Dunn (5) indicated that the autophagosomal membrane derives from the rough ER. However, the post-Golgi membrane, as well as a unique de novo synthesized membrane, the phagophore, have also been proposed as sources (6, 7). In recent morphological studies on yeast autophagy (8, 9), autophagosomal membranes were found to have features distinct from th...

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“…2) is catalyzed by betaine-homocysteine methyltransferase found in the liver, probably the kidney, and possibly in other tissues such as the brain [13]. In this reaction, gly…”

Section: Betaine Homeostasis and Metabolismmentioning

confidence: 99%