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Biomedical subjects

C Y Lai

Publications and source records attributed to C Y Lai.

At least 73 records · Page 4Linked to original sources
PubMed1984

The primary structure of the COOH-terminal half of cholera toxin subunit A1 containing the ADP-ribosylation site.

The sequence of 96 amino acid residues from the COOH-terminus of the active subunit of cholera toxin, A1, has been determined as (sequence; see text) This is the largest fragment obtained by BrCN cleavage of the subunit A1 (Mr 23,000), and has previously been indicated to contain the active site for the adenylate cyclase-stimulating activity. Unequivocal identification of the COOH-terminal structure was achieved by separation and analysis of the terminal peptide after the specific chemical cleavage at the only cysteine residue in A1 polypeptide. The site of self ADP-ribosylation in the A1 subunit [C. Y. Lai, Q.-C. Xia, and P.T. Salotra (1983) Biochem. Biophys. Res. Commun. 116, 341-348] has now been identified as Arg-50 of this peptide, 46 residues removed from the COOH-terminus. The cysteine that forms disulfide bridge to A2 subunit in the holotoxin is at position 91.

Adenosine Diphosphate Ribose↗
PubMed1984

Purification and properties of a toxin from the South African sea anemone, Pseudactinia varia.

A comparison was made of the hemolytic potency of aqueous extracts prepared from five species of intertidal sea anemones from the coast of South Africa. The active agent in an extract of Pseudactinia varia was purified by ammonium sulfate precipitation, gel permeation chromatography and isoelectric focusing. The hemolytic toxin, termed variolysin, is a protein having a molecular weight of 19,500 and an isoelectric pH of 9.8. It retained appreciable activity after heating to 70 degrees for 40 min. Amino acid analysis revealed that it lacked methionine and cysteine. Its hemolytic activity was inhibited by sphingomyelin. The properties of variolysin show that it is broadly similar to cytolytic toxins isolated from a number of other anthozoans.

Amino Acids↗
PubMed1983

Location and amino acid sequence around the ADP-ribosylation site in the cholera toxin active subunit A1.

Renatured, S-carboxymethylated subunit A1 of cholera toxin possess the ADP-ribose transferase activity (Lai, et.al., Biochem. Biophys. Res. Commun. 1981, 102, 1021). In the absence of acceptor self ADP-ribosylation of A1 subunit was observed. Stoicheometric incorporation of ADP-ribose moiety was achieved in 20 min at room temperature in a 0.1 - 0.2M PO4(Na) buffer, pH 6.6. On incubation of the complex with polyarginine, 75% of the enzyme-bound ADP-ribose moiety was transferred to the acceptor in 25 min. The ADP-ribosylated A1 was stable at low pH, and on cleavage with BrCN, the ADP-ribose moiety was found associated with peptide Cn I, the COOH-terminal fragment of A1 subunit. On further fragmentation with cathepsin D, a dodecapeptide containing ADP-ribose moiety was isolated whose structure was determined as: Asp-Glu-Glu-Leu-His-Arg-Gly-Tyr-Arg*-Asp-Arg-Tyr. The Arg* in the peptide was indicated to be the site of ADP-ribosylation.

Amino Acid Sequence↗
PubMed1983

Rabbit pulmonary angiotensin-converting enzyme: the NH2-terminal fragment with enzymatic activity and its formation from the native enzyme by NH4OH treatment.

The NH2-terminal sequence of 22 residues of rabbit lung angiotensin-converting enzyme has been determined as (NH2)Thr-Leu-Asp-Pro-Gly-Leu-Leu-Pro-Gly-Asp-Phe-Ala -Ala-Asp-Asn-Ala-Gly-Ala-Arg-Leu-Phe-Ala-. In the course of purification of the enzyme for structural analysis a protein of Mr = 82,000 with angiotensin-converting activity was separated from the major fraction containing the native enzyme (Mr = 140,000). This low-molecular-weight enzyme catalyzed the hydrolysis of the synthetic substrate Hip-His-Leu at a rate 23% of that with the native enzyme, and exhibited a similar Km value as well as behaviors towards various effectors of angiotensin-converting enzyme. Edman degradation of both the native and the 82K enzymes revealed that they contain identical amino acid sequences from the NH2-termini. This result and those of peptide mapping and carbohydrate and amino acid analyses indicate that the 82K enzyme is a fragment derived from the NH2-terminal portion of the native enzyme, and hence contains its catalytic site. Evidence has been obtained indicating that the active fragment was formed from the native enzyme during its elution from the antibody-affinity column with NH4OH: on treatment of the native enzyme (140K Mr) with 1 N NH4OH at room temperature, a cleavage occurred and two proteins with Mr = 82K and Mr = 62K were obtained. The 82K Mr fragment was found to be enzymatically active and to contain the same NH2-terminal sequence as the native enzyme. The other fragment (62K Mr) was devoid of the activity and was shown to derive from the COOH-terminal portion of the native enzyme by the peptide mapping and terminal analyses. Cleavage of a peptide bond with NH4OH is unusual and appears to be specific for the native angiotensin-converting enzyme from rabbit lung.

Amino Acids↗
PubMed1983

The role of protease in streptolysin S formation.

Production of streptolysin S by streptococci was found to be inhibited by treatment with protease inhibitors, tosylphenylalanine chloromethyl ketone (TPCK), tosyllysine chloromethyl ketone (TLCK), or phenylmethylsulfonyl fluoride (PMSF), even in the presence of the inducer oligonucleotides. Other protease inhibitors, antipain, leupeptin, or pepstatin were found to have little or no effect. Trypsin reversed the effect of TPCK or TLCK. The reversal was dependent upon the amount of added trypsin and the incubation time at 37 degrees C, suggesting that a protease activity was involved in the hemolysin formation. The effect of trypsin was not observed if chloramphenicol was also added, suggesting that a precursor of streptolysin S was processed as it was synthesized and released into medium as the active hemolysin, by the concerted action of a protease and inducer oligonucleotides. Experiments with the subcellular fractions of streptococci indicated that the streptolysin precursor was localized in the insoluble fraction and the "processing" protease in the supernatant fraction.

Cell-Free System↗
PubMed1981

Purification and characterization of recombinant human leukocyte interferon (IFLrA) with monoclonal antibodies.

Recombinant human leukocyte interferon produced in bacteria (IFLrA) was purified to homogeneity with the use of monoclonal antibodies against leukocyte interferon. The purified interferon exhibited a single band of Mr = approximately 19,000 on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Amino acid analysis and the NH2-terminal sequence were consistent with the sequence predicted from the DNA. Some of the purified product contained NH2-terminal methionine; the terminal methionine was removed from the rest of the chains.

Amino Acids↗
PubMed1981

Amino acid sequence of a human leukocyte interferon.

The primary structures of three major species of human leukocyte interferon differ from the structure predicted from the DNA sequence of recombinants containing leukocyte interferon-coding regions. Compared to the recombinant interferon produced in bacteria, three of the purified natural proteins isolated from leukocytes lack the 10 COOH-terminal amino acids suggested by the DNA sequence.

Amino Acid Sequence↗