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Biomedical subjects

Y T Li

Publications and source records attributed to Y T Li.

At least 55 records · Page 3Linked to original sources
PubMed1999

Comparison of the frequency-doubled Q-switched Nd:YAG laser and 35% trichloroacetic acid for the treatment of face lentigines.

BACKGROUND: Face lentigines are one of the manifestations of photodamaged skin and often put people in socially embarrassing situations. Several lasers have been used to remove lentigines at vast expense. However, trichloroacetic acid (TCA) is an alternative for treating lentigines that costs much less. OBJECTIVE: To compare the efficacy of the frequency-doubled Q-Switched Nd:YAG laser (532 nm) and 35% TCA for the treatment of face lentigines. METHODS: Twenty patients (Fitzpatrick skin Types III-IV) with a total of 37 lentigines on faces were randomly collected; each lentigo was divided into medial and lateral halves. Frequency-doubled Q-switched Nd:YAG laser (532 nm) and 35% TCA were applied to the medial and lateral halves of each lentigo respectively. The efficacy after 1 treatment was compared after 6 months. RESULTS: The frequency-doubled Q-switched Nd:YAG laser (532 nm) had a better result than that of 35% TCA for the treatment of facial lentigines. CONCLUSION: In order to get a better result after one treatment, the authors suggest the frequency-doubled Q-switched Nd:YAG laser (532 nm) rather than 35% TCA for treating lentigines even though the cost of frequency-doubled Q-switched Nd:YAG laser (532 nm) is greater than that of 35% TCA for both physician and patient.

Adult↗
PubMed1999

Structural characterization of gangliosides isolated from mullet milt using electrospray ionization-tandem mass spectrometry.

Electrospray ionization (ESI) coupled with tandem mass spectrometry has been used in conjunction with microwave-mediated saponification, periodate oxidation, and clostridial sialidase hydrolysis to enable detailed structural characterization of gangliosides and their derivatives present in mullet milt. The gangliosides extracted from mullet milt were determined to be GM3, GM3 lactone, GM3 methyl ester, and 9-O-acetyl GM3. For the major ganglioside GM3 and all GM3 derivatives, the ceramide composition was revealed to be C18:1/C16:0. GM3 with a C18:0/C16:0 ceramide was also found as a minor ganglioside. Both the ganglioside intramolecular ester and the ganglioside methyl ester (lacking carboxylic acid groups) showed dominant chloride attachment peaks (M + Cl)- in negative ion ESI-MS in addition to low intensity peaks corresponding to (M-H)-. GM3 and O-acetyl GM3 bearing carboxylic acid functions showed only (M-H)-. In positive ion ESI, GM3 and O-acetyl GM3 revealed (M + 2Na-H)+ peaks in addition to (M + Na)+, indicating free exchange of the carboxylic acid proton with a sodium cation, while the ganglioside intramolecular ester and ganglioside methyl ester with no acidic protons yielded only (M + Na)+. The strategy of employing ESI-MS to detect products of established wet chemical reactions represents a general approach for elucidation of ganglioside structural details.

Animals↗
PubMed1999

Psychosocial risk factors of teenage pregnancy in eastern Taiwan.

BACKGROUND: Teenage pregnancy is a growing worldwide problem, associated with adolescents' social and health problems and poor perinatal outcome. This study was conducted to investigate psychosocial factors predisposing to the high teenage pregnancy rate in Hualien, eastern Taiwan. METHODS: A regional hospital-based study was conducted with a retrospective analysis of hospital records and questionnaires to collect personal and family data regarding the perinatal outcome of 100 pregnant teenagers and 100 pregnant adults, who had normal deliveries at the Buddhist Tzu-Chi General Hospital between 1994 and 1995. RESULTS: A significantly higher percentage of teenage mothers were of aboriginal origin than adult mothers (63% vs 25%; p < 0.05) and lived in rural areas (80% vs 19%; p < 0.05). There was also a higher incidence of late antenatal care (31% vs 11%; p < 0.05) drinking (36% vs 9%; p < 0.05) and a greater history of smoking (34% vs 7%; p < 0.05) among teenage mothers. About 86% of teenage mothers did not use any contraception. The preterm birth rate was not significantly higher in the adult group, but teenage mothers tended to have significantly lower birth weight infants (19% vs 9%; p < 0.05) and a decreased incidence of cesarean section (19% vs 33%; p < 0.05). About 20% of the teenage mothers had their first coitus before the age of 13, while none of the adult mothers had sexual coitus before the age of 13. CONCLUSIONS: In this hospital-based study, teenage mothers tended to be of aboriginal origin, lived in rural areas, had early sexual exposure without contraception, had drinking and smoking habits, were late seeking antenatal care and gave birth to low birth weight infants.

Adolescent↗
PubMed1999

Susceptibility testing and clinical effect of fusidic acid in oxacillin-resistant Staphylococcus aureus infections.

One hundred and six oxacillin-resistant Staphylococcus aureus (ORSA) isolates collected from various clinical specimens at Mackay Memorial Hospital during the period from 1997 to 1998 were tested. The MICs of fusidic acid against ORSA isolates were in the range of 0.06 microgram/mL to 4 micrograms/mL. There was only one ORSA isolate (0.9%) resistant to fusidic acid in this study. Twenty-four patients with ORSA infection were enrolled into the study between July 1997 to June 1998. Four patients without the evidence of sepsis received oral fusidic acid only. The other 20 patients with sepsis received intravenous glycopeptide first, and then the oral fusidic acid for 7 to 10 days. Oral fusidic acid was used for mild ORSA infection and achieved satisfactory clinical outcome. In severe ORSA infection, it was found that sequential therapy with fusidic acid had relapsed in three patients. We concluded that oral fusidic acid achieved a satisfactory outcome in our patients with mild ORSA infection. In such cases, fusidic acid may provide an effective alternative treatment of choice. The rare side effect of granulocytopenia and jaundice was noted in our case series. The side effect may be not so rare as previously thought, if it had been carefully looked for.

Adult↗
PubMed1998

The crystal structure of an intramolecular trans-sialidase with a NeuAc alpha2-->3Gal specificity.

BACKGROUND: Intramolecular trans-sialidase from leech (Macrobdella decora) is a unique enzyme which cleaves the terminal neuraminic acid (NeuAc) residue from sialoglycoconjugates, releasing 2, 7-anhydro-neuraminic acid (2,7-anhydro-NeuAc). It is the first enzyme found to exhibit strictly specific cleavage of NeuAc alpha2-->3Gal linkages in sialoglycoconjugates. The release of 2,7-anhydro-NeuAc instead of NeuAc implies a unique mechanism, in which the sialosyl linkage is transferred within the sialoglycoconjugate rather than hydrolyzed. The aims of the structural study were to gain structural insight into the strict specificity and unique mechanism of this unusual enzyme. RESULTS: . The 2.0 A crystal structure of recombinant leech intramolecular trans-sialidase has been solved by multiple isomorphous replacement. The 1.8 A structure of the enzyme in complex with 2-deoxy-2, 3-didehydro-NeuAc was also solved. The refined model comprising residues 81-769 has a catalytic beta-propeller domain (C), a N-terminal lectin-like domain (II) and an irregular beta-stranded domain (III) inserted into the catalytic domain. The structure reveals several possible carbohydrate-binding features: domain II has a concave face, like that of other sialidases, and there is a suitable surface charge distribution at the domain III-C interface. CONCLUSIONS: Structural comparisons showed closer evolutionary relationships to bacterial sialidases than to viral neuraminidases. Mainchain and sidechain atoms around Thr593 make the glycerol-binding pocket incapable of accommodating an extended equatorial 6-glycerol group, implying that the 6-glycerol group of the reaction intermediate may occupy an axial position, which is also required by the catalytic mechanism. The steric hindrance introduced by the bulky sidechain of Trp734 above the 2-carboxylate group may explain the lack of water involvement in the cleavage reaction and the substrate specificity.

Amino Acid Sequence↗
PubMed1998

Antibodies to GT1a ganglioside in patients with Guillain-Barré syndrome.

Serum antibodies from 8 (13%) of 62 patients with the acute Guillain-Barré syndrome (GBS) and 1 of 3 patients with the Miller Fisher syndrome (MFS) recognized a minor ganglioside in bovine and human brain trisialoganglioside fractions. The ganglioside antigen migrated between GD1a and GD1b on thin-layer chromatograms. The structure of this ganglioside was established to be GT1a by thin-layer chromatography blotting and mass spectrometry. GT1a a ganglioside was also detected in human and bovine peripheral nerves by thin-layer chromatogram immunostaining. Serum from the GBS patients had IgM, IgG, or IgA antibodies against GT1a detectable by enzyme-linked immunosorbent assay (ELISA). Serum from the MFS patient also had elevated levels of IG against GT1a. None of the sera from 43 patients with other neurological diseases or from 24 healthy controls reacted with GT1a. Sera from 6 of 8 GBS patients with anti-Gt1a antibodies also reacted with GQ1b. There was no difference in the incidence of anti-GT1a immunoglobulins in acute GBS patients with or without oculomotor abnormalities. Levels of anti-GT1a antibodies correlated temporally wit clinical symptoms in GBS patients. Although the incidence of dysphagia was slightly higher in GBS patients with anti-GT1a antibodies than in those without, the number of patients studied may have been too small to detect an association between anti-GT1a antibodies and an a specific clinical variant of GBS. Our data demonstrate that a proportion of GBS patients have antibodies against GT1a ganglioside and suggest that these antibodies may play a role in the pathogenesis of neuropathy in GBS.

Adult↗
PubMed1998

Specificity of mouse GM2 activator protein and beta-N-acetylhexosaminidases A and B. Similarities and differences with their human counterparts in the catabolism of GM2.

Tay-Sachs disease, an inborn lysosomal disease featuring a buildup of GM2 in the brain, is caused by a deficiency of beta-hexosaminidase A (Hex A) or GM2 activator. Of the two human lysosomal Hex isozymes, only Hex A, not Hex B, cleaves GM2 in the presence of GM2 activator. In contrast, mouse Hex B has been reported to be more active than Hex A in cleaving GM2 (Burg, J., Banerjee, A., Conzelmann, E., and Sandhoff, K. (1983) Hoppe Seyler's Z. Physiol. Chem. 364, 821-829). In two independent studies, mice with the targeted disruption of the Hexa gene did not display the severe buildup of brain GM2 or the concomitant abnormal behavioral manifestations seen in human Tay-Sachs patients. The results of these two studies were suggested to be attributed to the reported GM2 degrading activity of mouse Hex B. To clarify the specificity of mouse Hex A and Hex B and to better understand the observed results of the mouse model of Tay-Sachs disease, we have purified mouse liver Hex A and Hex B and also prepared the recombinant mouse GM2 activator. Contrary to the findings of Burg et al., we found that the specificities of mouse Hex A and Hex B toward the catabolism of GM2 were not different from the corresponding human Hex isozymes. Mouse Hex A, but not Hex B, hydrolyzes GM2 in the presence of GM2 activator, whereas GM2 is refractory to mouse Hex B with or without GM2 activator. Importantly, we found that, in contrast to human GM2 activator, mouse GM2 activator could effectively stimulate the hydrolysis of GA2 by mouse Hex A and to a much lesser extent also by Hex B. These results provide clear evidence on the existence of an alternative pathway for GM2 catabolism in mice by converting GM2 to GA2 and subsequently to lactosylceramide. They also provide the explanation for the lack of excessive GM2 accumulation in the Hexa gene-disrupted mice.

Animals↗
PubMed1998

Synthesis of aryl 3'-sulfo-beta-lactosides as fluorogenic and chromogenic substrates for ceramide glycanases.

We have previously reported that 4-methylumbelliferyl 6'-O-benzyl-beta-lactoside (2) is a useful substrate for a fluorometric assay of ceramide glycanase (CGase) (L.-X. Wang, N. V. Pavlova, S.-C. Li, Y.-T. Li and Y. C. Lee, Glycoconjugate J., 13 (1996) 359-365). The introduction of a 6-O-benzyl group at the terminal Gal efficiently protected the substrate from its hydrolysis by exo-galactosidase, permitting the assay of CGase in crude biological materials. However, a drawback of this substrate is its low water-solubility and relatively high Km (at a mM level). Introduction of a sulfate group into 4-methylumbelliferyl beta-lactoside (1) led to the formation of 4-methylumbelliferyl 3'-O-sulfo-beta-lactoside (3), which was found to be a more effective substrate than 2. Moreover, the presence of a 3'-O-sulfate group not only increases the water solubility tremendously, but also protects the substrate from cleavage by exo-beta-galactosidase as the 6'-O-benzyl group in 2 does. In addition to the fluorogenic substrate (3), two sulfated chromogenic substrates, N-tetradecanoyl-4-O(3'-sulfo-beta-lactosyl)-3-nitro-L-tyrosine methyl ester (9) and 2-N-(tetradecanoylamino)-4-nitro-phenyl 3'-sulfo-beta-lactoside (12), were synthesized and their suitability for a photometric assay of CGase was evaluated. Substrates 9 and 12, with a long fatty acid chain attached to the aglycon part, have a Km value close to that of the natural substrate GM1 (at a microM level).

Chromogenic Compounds↗
PubMed1998

Diagnosis and treatment of adrenal tumours: a review of 35 years' experience.

OBJECTIVE: To review and analyse clinical data on the diagnosis and management of patients with adrenal masses. PATIENTS AND METHODS: Patients admitted with adrenal masses between 1960 and 1995 were reviewed. The series comprised 116 males (mean age 41.4 years, SD 10.5, range 3-77) and 95 females (mean 36.9 years, SD 11.6, range 1-62); eight patients were < 14 years old and the overall mean (SD) age was 39.4 (12.8) years. The diagnosis was based on symptoms, signs, hormone levels and imaging studies. All tumours were confirmed by surgery, and pathology and results of analysis assessed statistically. RESULTS: Over the last 35 years, the incidence of adrenal tumours was 1.7% of all patients admitted with genitourological diseases or 9.7% of patients with genitourinary tumours at our institution. The prevalence of adrenal tumours in males and females was similar but Cushing's syndrome was 3.1 times more frequent in females than in males and phaeochromocytoma 1.9 times more frequent in males than females. Most patients with adrenal tumour were aged 30-50 years. Of 211 adrenal tumours, 151 (72%) were functional, with a prevalence of benign tumour, and 60 (28%) were nonfunctional, with 35% malignant. There were 78 'incidentalomas' which included 18 functional tumours. Overall, 210 tumours were removed and one was explored. Correlation analysis between tumour size and character or hormone levels showed that size was significantly positively correlated with tumour character (r = 0.4010, P < 0.001), but there was no relationship between tumour size and hormonal levels. The postoperative complication rate was 3.3% and the mortality 0.5%. CONCLUSIONS: Based on this analysis we recommend that computed tomography is the first method used to define and localize the adrenal masses, that hormone levels should be determined in symptomatic or asymptomatic patients with adrenal masses, and that functional adrenal tumours and solid incidentalomas of any size should be removed surgically.

Adolescent↗
PubMed1997

alpha-KDOase activity in oyster and synthesis of alpha- and beta-4-methylumbelliferyl ketosides of 3-deoxy-D-manno-octulosonic acid.

Although alpha- and beta-linked 3-deoxy-D-manno-octulosonic acid (KDO) is found in lipopolysaccharides (LPSs) of Gram-negative bacteria, capsular polysaccharides of microorganisms, and plants, very little is known about its degradation. Using both thin-layer chromatography and the periodate-thiobarbituric acid reaction, we found that the hepatopancreas of oyster (Crassostrea virginica) contained an enzyme (alpha-KDOase) capable of releasing alpha-linked KDO from LPSs. To facilitate the studies of alpha-KDOase, we have carried out the synthesis of 4-methylumbelliferyl-alpha-KDO (alpha-KDO-MU) by conjugating the glycosyl chloride of the per-O-acetylated methylester of KDO with methylumbelliferone by the SN2 type reaction and the catalyzed phase-transfer. In both cases, the beta-anomer was obtained as the major product with a yield of about 80%, whereas the yield of alpha-anomer was only about 7%. Attempts to increase the yield of alpha-anomer were not successful. alpha-KDO-MU was used as substrate to follow the purification of alpha-KDOase from oyster hepatopancreas. The pH optimum for oyster alpha-KDOase was determined to be 4.5 using Re-LPS as substrate and 3.0 using alpha-KDO-MU as substrate. The enzyme was found to be stable in the pH range of 3-8. This enzyme released KDO from different LPSs, including Re-LPS from Escherichia coli and Salmonella minnesota, Rd-LPS from S. minnesota, and de-O-acyl-Re-LPS (Kiang, J., Szu, S. C., Wang, L.X., Tang, M., and Lee, Y. C. (1997) Anal. Biochem. 245, 97-101).

Animals↗
PubMed1997

Hippocampus in delay eyeblink classical conditioning: essential for nefiracetam amelioration of learning in older rabbits.

Rabbits acquire conditioned responses (CRs) normally with bilateral removal of the hippocampus, but alterations of the intact hippocampus can affect the rate of acquisition. The cognition-enhancing drug, nefiracetam ameliorated the acquisition of CRs in older rabbits, protected membrane dysfunction in hippocampal CA1 neurons following oxygen and glucose deprivation, and promoted the release of diverse neurotransmitters, including acetylcholine. Because the septo-hippocampal cholinergic system is demonstrated to be involved in eyeblink conditioning, this experiment was undertaken to explore whether nefiracetam ameliorates conditioning via the hippocampus. Data from 53 rabbits of a mean age of 28 months were tested under two drug conditions (10 or 0 mg/kg nefiracetam) and 4 lesion conditions (bilateral hippocampectomy, bilateral neocortical removal, sham surgery, no surgery). The three groups of nefiracetam-treated rabbits with intact hippocampus acquired CRs more rapidly than the vehicle-treated groups, but rabbits with bilateral hippocampectomy treated with nefiracetam learned like vehicle-treated rabbits. Results suggest that nefiracetam ameliorates learning via the hippocampus. Because of the parallels between conditioning in rabbits with disrupted hippocampal cholinergic systems and conditioning in Alzheimer's disease (AD), these results suggest that nefiracetam may ameliorate conditioning in AD as it ameliorates conditioning in older rabbits.

Aging↗
PubMed1997

Interaction of GM2 activator protein with glycosphingolipids.

GM2 activator protein is a protein cofactor that has been shown to stimulate the enzymatic hydrolysis of both GalNAc and NeuAc from GM2 (Wu, Y. Y., Lockyer, J. M., Sugiyama, E., Pavlova, N.V., Li, Y.-T., and Li, S.-C. (1994) J. Biol. Chem. 269, 16276-16283). To understand the mechanism by which GM2 activator stimulates the hydrolysis of GM2, we examined the interaction of this activator protein with GM2 as well as with other glycosphingolipids by TLC overlay and Sephacryl S-200 gel filtration. The TLC overlay analysis unveiled the binding specificity of GM2 activator, which was not previously revealed. Under the conditions optimal for the activator protein to stimulate the hydrolysis of GM2 by beta-hexosaminidase A, GM2 activator was found to bind avidly to acidic glycosphingolipids, including gangliosides and sulfated glycosphingolipids, but not to neutral glycosphingolipids. The gangliosides devoid of sialic acids, such as asialo-GM1 and asialo-GM2, and the GM2 derivatives whose carboxyl function in the NeuAc had been modified by methyl esterification or reduction, were only very weakly bound to GM2 activator. These results indicate that the negatively charged sugar residue or sulfate group in gangliosides is one of the important sites recognized by GM2 activator. For comparison, we also studied in parallel the complex formation between glycosphingolipids and saposin B, a separate activator protein with broad specificity to stimulate the hydrolysis of various glycosphingolipids. We found that saposin B bound to neutral glycosphingolipids and gangliosides equally well, and there was an exceptionally strong binding to sulfatide. In contrast to previous reports, we found that GM2 activator formed complexes with GM2 and other gangliosides in different proportions depending on the ratio between the activator protein and the ganglioside in the incubation mixture prior to gel filtration. We were not able to detect the specific binding of GM2 activator to GM2 when GM2 was mixed with GM1 or GM3. Thus, the specificity or the mode of action of GM2 activator cannot be simply explained by its interaction with glycosphingolipids based on complex formation. The binding of GM2 activator to a wide variety of negatively charged glycosphingolipids may indicate that this activator protein has functions other than assisting the enzymatic hydrolysis of GM2.

Brain Chemistry↗
PubMed1997

Quantitative determination of Orlistat (tetrahydrolipostatin, Ro 18-0647) in human plasma by high-performance liquid chromatography coupled with ion spray tandem mass spectrometry.

A rapid, sensitive and specific analytical method was developed and validated to quantify tetrahydrolipostatin (Orlistat, Ro 18-0647) in human plasma in order to provide pharmacokinetic data from clinical trials. This method employs a preliminary plasma protein precipitation step followed by a simple, one-step liquid-liquid extraction procedure to isolate Ro 18-0647 and its pentadeuterated internal standard, Ro 18-0647-d5, from the biological matrix. Reconstituted extracts were analyzed by liquid chromatography/ion spray tandem mass spectrometry (LC/MS/MS) in the selected reaction monitoring (SRM) mode. Chromatography was carried out using a 2 mm i.d. x 50 mm Deltabond Phenyl column. The eluent was acetonitrile-2 mM ammonium acetate (90:10). The retention time of the analyte was 1.2 min and chromatographic run times were less than 1.5 min. No interferences from anticoagulants, collection devices or endogenous constituents of the plasma were observed. The assay has a lower limit of quantitation (LLQ) of 0.20 ng ml-1 in plasma and a lower limit of detection (LLD) of 0.10 ng ml-1 plasma, based on 1 ml aliquots. The capability to detect 0.025 ng ml-1 in plasma has also been demonstrated. The calibration graphs were linear from 0.20 to 10 ng ml-1. The assay was initially validated with a linear range of 0.20-1.0 ng ml-1. This range was later extended and validated to an upper level of quantitation of 10 ng ml-1. Intra- and inter-assay precision studies showed a mean variability of less than 10%. The recovery, inter-assay precision and accuracy of the method were within acceptable bioanalytical standards. The assay has been shown to reliably provide automated, unattended sample analysis for approximately 150 samples per day. In an additional series of tests, Ro 18-0467 was shown to be stable under conditions that might be encountered during the analysis of samples from clinical trials. This LC/MS/MS assay procedure for Ro 18-0647 in human plasma has proven to be robust, sensitive, specific, accurate and reproducible. This method has been used to analyze over 5000 study samples.

Calibration↗
PubMed1997

Structural organization and expression of the gene for the mouse GM2 activator protein.

The GM2 activator protein is an essential component for the degradation of GM2 ganglioside by hexosaminidase A in vivo. Mutations in the human gene coding for the GM2 activator protein cause the AB variant of GM2-gangliosidosis, a condition that is clinically indistinguishable from Tay-Sachs disease. To understand better factors affecting the expression of the GM2 activator protein gene (Gm2a) in mouse tissues, we have determined its exon-intron organization and analyzed its promoter region. Gm2a is about 14 kb, has four exons, and the 5' flanking region contains a CAAT box, Sp1 binding sites, AP-1, AP-2 sites, and a pair of IRE sites. A 1.2-kb fragment upstream from the initiation codon was shown to have promoter activity in NIH 3T3 cells. Similarities between the elements present in Gm2a and Hexa promoters might in part explain their similar expression patterns in mouse tissues. The different levels of GM2 activator protein mRNA in liver, kidney, brain, and testis are not owing to the use of different transcription start sites, because a single start site was found 50 bp upstream from the initiation codon in each these tissues. Northern blot analysis demonstrated variation in the GM2 activator protein mRNA expression during mouse development. Gm2a was mapped to Chromosome (Chr) 11, where it co-segregated with Csfgm.

3T3 Cells↗
PubMed1996

Cloning and expression of sialidase L, a NeuAcalpha2-->3Gal-specific sialidase from the leech, Macrobdella decora.

Sialidase L is a NeuAcalpha2-->3Gal linkage-specific sialidase that releases 2,7-anhydro-NeuAc instead of NeuAc from sialoglycoconjugates (Chou, M.-Y., Li, S.-C., Kiso, M., Hasegawa, A., and Li, Y.-T.(1994) J. Biol. Chem. 269, 18821-18826). A 2. 5-kilobase cDNA of sialidase L was cloned by a combination of methods based on polymerase chain reactions. The composite cDNA sequence reveals an open reading frame coding for 762 amino acids, including a putative 28-residue signal peptide at the N terminus that is similar to the signal sequence of the Clostridium septicum sialidase. The result suggests that sialidase L is a secretory enzyme. The coding sequence excluding the putative signal peptide of sialidase L was overexpressed in Escherichia coli. The purified recombinant enzyme was characterized to be as active as the enzyme isolated from the leech. It also possessed the strict NeuAcalpha2-->3Gal linkage specificity and released the unique cleavage product, 2,7-anhydro-NeuAc from sialoglycoconjugates. The deduced amino acid sequence of sialidase L exhibits little similarity with other reported sialidases. However, sialidase L contains a conserved "FRIP region" and four repeating "Asp box" motifs that align well with the corresponding positions of bacterial sialidases. The predicted beta-strand structures near the conserved motifs of sialidase L are similar to those of Salmonella typhimurium sialidase. Several conserved single amino acid residues of bacterial sialidases, including those known to be involved in the active site of Salmonella enzyme, are conserved in the deduced amino acid sequence of sialidase L. This observation suggests that part of the catalytic mechanism of sialidase L may be similar to the ordinary sialidase.

Amino Acid Sequence↗
PubMed1996

Characterization of an alternatively spliced GM2 activator protein, GM2A protein. An activator protein which stimulates the enzymatic hydrolysis of N-acetylneuraminic acid, but not N-acetylgalactosamine, from GM2.

GM2 activator protein is a protein cofactor which stimulates the enzymatic hydrolysis of both GalNAc and NeuAc from GM2. We have previously isolated two cDNA clones, GM2 activator cDNA and GM2A cDNA, for human GM2 activator protein (Nagarajan, S., Chen, H.-C., Li, S.-C., Li, Y.-T., and Lockyer, J. M. (1992) Biochem. J. 282, 807-813). GM2A mRNA is an RNA alternative splicing product that contains exons 1, 2, 3, and intron 3 of the genomic DNA sequence of GM2 activator protein (Klima, H., Tanaka, A., Schnabel, D., Nakano, T., Schröder, M., Suzuki, K., and Sandhoff, K. (1991) FEBS Lett. 289, 260-264). GM2A cDNA encodes a protein (GM2A protein) containing 1-109 of the 160 amino acids of human GM2 activator protein, plus a tripeptide (VST) encoded by intron 3 at the COOH terminus. Thus, GM2A protein can be regarded as a form (truncated version) of GM2 activator protein. We have expressed GM2A cDNA in Escherichia coli using pT7-7 as the vector. The recombinant GM2A protein was purified to an electrophoretically homogeneous form and was found to stimulate the hydrolysis of NeuAc from GM2 by clostridial sialidase, but not the hydrolysis of GalNAc from GM2 by beta-hexosaminidase A. Like GM2 activator protein, GM2A protein also specifically recognized the terminal GM2 epitope in GalNAc-GD1a and stimulated the hydrolysis of only the external NeuAc from this ganglioside by clostridial sialidase. These results enabled us to discern the enzymatic hydrolyses of GalNAc and NeuAc from the GM2 epitope and established that the NeuAc recognition domain of GM2 activator protein is located within amino acids 1-109. The presence of GM2A mRNA in human tissues and the selective stimulation of NeuAc hydrolysis by GM2A protein indicate that this activator protein may be involved in the catabolism of GM2 through the asialo-GM2 pathway.

Acetylgalactosamine↗