scholarly journals Isolation and sequence of a cDNA for human pro-(cathepsin L)

1988 ◽  
Vol 253 (1) ◽  
pp. 303-306 ◽  
Author(s):  
S Gal ◽  
M M Gottesman
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Amino Acid Sequence Identity ◽  
Cathepsin L ◽  
Acid Proteinase ◽  
Cdna Clones ◽  
Terminal Sequence ◽  
Human Form ◽  
Cross Hybridization ◽  
Human Cathepsin

The major excreted protein (MEP) of malignantly transformed mouse fibroblasts is the precursor to an acid proteinase with enzymic specificity similar to that of human cathepsin L. By cross-hybridization with a mouse MEP sequence, cDNA clones of the human form of MEP in an SV40 expression vector were isolated. A 1.6 kb cDNA showed 70% deduced amino acid sequence identity with mouse MEP. The deduced amino acid sequence of the cloned human MEP was the same, except for two amino acids, as the N-terminal sequence of mature human cathepsin L, thereby establishing that human MEP is human pro-(cathepsin L). Use of this human pro-(cathepsin L) cDNA clone allowed the detection of a 1.6-1.8 kb pro-(cathepsin L) mRNA in human cells which was not detected with a mouse pro-(cathepsin L) probe.

scholarly journals The primary structure of the VLA-2/collagen receptor alpha 2 subunit (platelet GPIa): homology to other integrins and the presence of a possible collagen-binding domain.

1989 ◽  
Vol 109 (1) ◽  
pp. 397-407 ◽  
Author(s):  
Y Takada ◽  
M E Hemler
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Matrix Protein ◽  
Transmembrane Domain ◽  
Cdna Clones ◽  
Terminal Sequence ◽  
Collagen Binding ◽  
Collagen Receptor ◽  
Integrin Family

VLA-2 (also called gpIa/IIa on platelets) is a collagen receptor with a unique alpha subunit and a beta subunit common to other adhesion receptors in the VLA/integrin family. Multiple cDNA clones for the human VLA-2 alpha 2 subunit have been selected from a lambda gtll library by specific antibody screening. The 5,374-bp nucleotide sequence encoded for 1,181 amino acids, including a signal peptide of 29 amino acids followed by a long extracellular domain (1,103 amino acids), a transmembrane domain, and a short cytoplasmic segment (22 amino acids). Direct sequencing of purified alpha 2 protein confirmed the identity of the 15 NH2-terminal amino acids. Overall, the alpha 2 amino acid sequence was 18-25% similar to the sequences known for other integrin alpha subunits. In particular, the alpha 2 sequence matched other integrin alpha chains in (a) the positions of 17 of its 20 cysteine residues; (b) the presence of three metal-binding domains of the general structure DXDXDGXXD; and (c) the transmembrane domain sequence. In addition, the alpha 2 sequence has a 191-amino acid insert (called the I-domain), previously found only in leukocyte integrins of the beta 2 integrin family. The alpha 2 I-domain was 23-41% similar to domains in cartilage matrix protein and von Willebrand factor, which are perhaps associated with collagen binding. The NH2-terminal sequence reported here for alpha 2 does not match the previously reported alpha 2 NH2-terminal sequence (Takada, Y., J. L. Strominger, and M. E. Hemler. 1987. Proc. Natl. Acad. Sci. USA. 84:3239-3243). Resolution of this discrepancy suggests that there may be another VLA heterodimer that resembles VLA-2 in size but has a different amino acid sequence.

Characterization of a novel heterodimeric cathepsin L-like protease and cDNA encoding the catalytic subunit of the protease in embryos of Artemia franciscana

2001 ◽  
Vol 79 (1) ◽  
pp. 43-56 ◽  
Author(s):  
Ann M Butler ◽  
Andrea L Aiton ◽  
Alden H Warner
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Amino Acid Sequence Identity ◽  
Single Gene ◽  
Cathepsin L ◽  
Cysteine Proteases ◽  
Artemia Franciscana ◽  
Catalytic Subunit ◽  
Mature Protein ◽  
Sequence Identity

Embryos and larvae of the brine shrimp, Artemia franciscana, contain a novel cathepsin L-like cysteine protease (ACP) composed of 28.5- and 31.5-kDa subunits. Both subunits of the ACP are glycosylated, and seven isoforms of the protease were identified by isoelectric focusing with pI values ranging from 4.6 to 6.2. Several clones containing sequences coding for the 28.5-kDa subunit of the ACP were isolated from an Artemia embryo cDNA library in lambda ZAP II. One clone of 1229 bp, with an open reading frame of 1014 bp, was sequenced and found to contain 50-65% amino acid sequence identity with several members of the cathepsin L subfamily of cysteine proteases. The mature protein predicted from this sequence consisted of 217 amino acids with a mass of 23.5 kDa prior to post-translational modifications. The mature protein showed 68.6% amino acid sequence identity with human cathepsin L and 73.9% identity with cathepsin L-like proteases from Sarcophaga. peregrina and Drosophila melanogaster. The full-length cDNA clone analyzed in this study (pCP-3b) was renamed AFCATL1 (A. franciscana Cathepsin L1) and the sequence has been deposited in the Genbank database, accession number AF147207. Northern blot analyses identified a single transcript of about 1.4 kb in both embryos and young larvae of Artemia. Southern blot analyses of Artemia genomic DNA treated with various restriction endonucleases indicated a single gene for the ACP. The catalytic subunit of the ACP was tightly associated with a 31.5-kDa protein, which may localize the protease to nonlysosomal sites in embryos and larvae.Key words: cathepsin L, proteases, embryos, development, Artemia.

C-Terminal amino acid sequence of chicken pepsinogen and its homology with sequences of other acid proteases

1980 ◽  
Vol 45 (4) ◽  
pp. 1144-1154 ◽  
Author(s):  
Miroslav Baudyš ◽  
Helena Keilová ◽  
Vladimír Kostka
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
The Other ◽  
Terminal Sequence ◽  
Terminal Part ◽  
Terminal Amino Acid ◽  
Tryptic Peptides ◽  
Terminal Amino ◽  
High Degree ◽  
Terminal Amino Acid Sequence

To determine the primary structure of the C-terminal part of the molecule of chicken pepsinogen the tryptic, chymotryptic and thermolytic digest of the protein were investigated and peptides derived from this region were sought. These peptides permitted the following 21-residue C-terminal sequence to be determined: ...Ile-Arg-Glu-Tyr-Tyr-Val-Ile-Phe-Asp-Arg-Ala-Asn-Asn-Lys-Val-Gly-Leu-Ser-Pro-Leu-Ser.COOH. A comparison of this structure with the C-terminal sequential regions of the other acid proteases shows a high degree of homology between chicken pepsinogen and these proteases (e.g., the degree of homology with respect to hog pepsinogen and calf prochymosin is about 66%). Additional tryptic peptides, derived from the N-terminal part of the zymogen molecule whose amino acid sequence has been reported before, were also obtained in this study. This sequence was extended by two residues using an overlapping peptide. An ancillary result of this study was the isolation of tryptic peptides derived from other regions of the zymogen molecule.

scholarly journals Hyaluronan-binding region of aggrecan from pig laryngeal cartilage. Amino acid sequence, analysis of N-linked oligosaccharides and location of the keratan sulphate

1992 ◽  
Vol 286 (3) ◽  
pp. 761-769 ◽  
Author(s):  
F P Barry ◽  
J U Gaw ◽  
C N Young ◽  
P J Neame
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Signal Peptidase ◽  
Amino Acid Residues ◽  
Terminal Sequence ◽  
Binding Region ◽  
Keratan Sulphate ◽  
Laryngeal Cartilage ◽  
Hyaluronan Binding

The hyaluronan-binding region (HABR) was prepared from pig laryngeal cartilage aggrecan and the amino acid sequence was determined. The HABR had two N-termini: one N-terminal sequence was Val-Glu-Val-Ser-Glu-Pro (367 amino acids in total), and a second N-terminal sequence (Ala-Ile-Ser-Val-Glu-Val; 370 amino acids in total) was found to arise due to alternate cleavage by the signal peptidase. The N-linked oligosaccharides were analysed by examining their reactivity with a series of lectins. It was found that the N-linked oligosaccharide on loop A was of the mannose type, while that on loop B was of the complex type. No reactivity was detected between the N-linked oligosaccharide on loop B' and any of the lectins. The location of keratan sulphate (KS) in the HABR was determined by Edman degradation of the immobilized KS-containing peptide. The released amino acid derivatives were collected and tested for the presence of epitope to antibody 5-D-4. On the basis of 5-D-4 reactivity and sequencing yields, the KS chains are attached to threonine residues 352 and 357. There is no KS at threonine-355. This site is not in fact in G1, but about 16 amino acid residues into the interglobular domain. Comparison of the structure of the KS chain from the HABR and from the KS domain of pig laryngeal cartilage aggrecan was made by separation on polyacrylamide gels of the oligosaccharides arising from digestion with keratanase. Comparison of the oligosaccharide maps suggests that the KS chains from both parts of the aggrecan molecule have the same structure.

scholarly journals Cloning, sequencing and heterologous expression of a cDNA encoding pigeon liver carnitine acetyltransferase

1995 ◽  
Vol 305 (2) ◽  
pp. 439-444 ◽  
Author(s):  
T M Johnson ◽  
H P Kocher ◽  
R C Anderson ◽  
G M Nemecek
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Heterologous Expression ◽  
Stop Codon ◽  
Expression System ◽  
Breast Muscle ◽  
Cdna Clones ◽  
Carnitine Acetyltransferase ◽  
Western Blots ◽  
Pigeon Liver

Two overlapping cDNA clones encoding pigeon liver carnitine acetyltransferase (EC 2.3.1.7) (CAT) were isolated from a pigeon liver lambda gt11 cDNA library by gene amplification using oligonucleotide primers based on the N-terminal amino acid sequence of the enzyme. The two clones, which represent the 5′ and 3′ ends of the gene, were spliced together to form a single cDNA construct containing the entire coding sequence for CAT, with an in-frame TGA stop codon 42 bases before the first ATG start site and a 3′-untranslated segment of 1057 bases. The largest open reading frame of 1942 nucleotides predicted a polypeptide of 627 amino acids and a molecular mass of 71.1 kDa. The N-terminus and four internal peptides from the amino acid sequence of pigeon breast muscle CAT were identified in the predicted sequence of the liver cDNA clone. The identity of the CAT cDNA was confirmed by heterologous expression of active recombinant CAT (rCAT) in insect cells using the baculovirus expression system. Western blots of rCAT from infected insect cell lysates and immunodetection with a rabbit anti-CAT polyclonal serum showed an immunoreactive protein band similar in size to native CAT from pigeon breast muscle. Like the native enzyme, rCAT was capable of acylating carnitine with a preference for small-chain acyl-CoAs of carbon chain lengths C2-C4.

scholarly journals Biochemical characterization of a second family of human Ia molecules, HLA-DS, equivalent to murine I-A subregion molecules.

1982 ◽  
Vol 156 (2) ◽  
pp. 550-566 ◽  
Author(s):  
S M Goyert ◽  
J E Shively ◽  
J Silver
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Amino Acid Sequence Identity ◽  
Biochemical Characterization ◽  
Amino Acid Sequence Homology ◽  
Molecular Weights ◽  
Hla Dr ◽  
D Region ◽  
Polypeptide Chains

In mice, two families of structurally distinct Ia molecules, one designated I-A and the other I-E, have been identified and characterized. The HLA-DR molecules represent one family of human Ia molecules equivalent to the murine I-E molecules on the basis of amino acid sequence homology. We describe the isolation and biochemical characterization of a second family of human Ia molecules, designated HLA-DS for second D-region locus, equivalent to the murine I-A molecules. The human HLA-DS molecules consist of two polypeptide chains, DS alpha (37,000 mol wt) and DS beta (29,000 mol wt), with 73% amino acid sequence identity to the murine I-A molecules. Furthermore, the HLA-DS molecules are closely linked genetically to HLA-DR molecules, a situation analogous to that observed in mice. The similarity in molecular weights of the DR and DS molecules might explain why others have failed to identify the latter in man.

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