Isolation and biochemical characterization of LEAP‐2, a novel blood peptide expressed in the liver

The human genome contains numerous genes whose protein products are unknown in terms of structure, interaction partner, expression, and function. To unravel the function of these orphan genes, it is of particular value to isolate native forms of protein and peptide products derived from these genes....

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Published in:	Protein science Vol. 12; no. 1; pp. 143 - 152
Main Authors:	Krause, Alexander, Sillard, Rannar, Kleemeier, Burkhard, Klüver, Enno, Maronde, Erik, Conejo‐García, José Ramon, Forssmann, Wolf Georg, Schulz‐Knappe, Peter, Nehls, Michael C., Wattler, Frank, Wattler, Sigrid, Adermann, Knut
Format:	Journal Article
Language:	English
Published:	Bristol Cold Spring Harbor Laboratory Press 01-01-2003
Subjects:	Alternative Splicing Amino Acid Sequence Animals Anti-Infective Agents - chemistry Anti-Infective Agents - isolation & purification Anti-Infective Agents - pharmacology antimicrobial activity Antimicrobial Cationic Peptides - chemistry Antimicrobial Cationic Peptides - genetics Antimicrobial Cationic Peptides - isolation & purification Antimicrobial Cationic Peptides - pharmacology Base Sequence Blood Proteins - chemistry Blood Proteins - genetics Blood Proteins - isolation & purification Blood Proteins - pharmacology CFU, colony‐forming unit Cloning, Molecular disulfide bonds Disulfides - chemistry DNA, Complementary - genetics Dose-Response Relationship, Drug ESIMS, electrospray ionization mass spectrometry hemofiltrate Hemofiltration Humans LEAP, liver‐expressed antimicrobial peptide liver Liver - metabolism Medicin och hälsovetenskap Microbial Sensitivity Tests Molecular Sequence Data Organ Specificity peptide RT‐PCR, reverse transcription/polymerase chain reaction Saccharomyces cerevisiae - drug effects secretion Sequence Alignment Spectrometry, Mass, Electrospray Ionization TFA, trifluoroacetic acid
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Summary:	The human genome contains numerous genes whose protein products are unknown in terms of structure, interaction partner, expression, and function. To unravel the function of these orphan genes, it is of particular value to isolate native forms of protein and peptide products derived from these genes. From human blood ultrafiltrate, we characterized a novel gene‐encoded, cysteine‐rich, and cationic peptide that we termed liver‐expressed antimicrobial peptide 2 (LEAP‐2). We identified several circulating forms of LEAP‐2 differing in their amino‐terminal length, all containing a core structure with two disulfide bonds formed by cysteine residues in relative 1–3 and 2–4 positions. Molecular cloning of the cDNA showed that LEAP‐2 is synthesized as a 77‐residue precursor, which is predominantly expressed in the liver and highly conserved among mammals. This makes it a unique peptide that does not exhibit similarity with any known human peptide regarding its primary structure, disulfide motif, and expression. Analysis of the LEAP‐2 gene resulted in the identification of an alternative promoter and at least four different splicing variants, with the two dominating transcripts being tissue‐specifically expressed. The largest native LEAP‐2 form of 40 amino acid residues is generated from the precursor at a putative cleavage site for a furin‐like endoprotease. In contrast to smaller LEAP‐2 variants, this peptide exhibited dose‐dependent antimicrobial activity against selected microbial model organisms. LEAP‐2 shares some characteristic properties with classic peptide hormones and it is expected that the isolation of this novel peptide will help to unravel its physiological role.
Bibliography:	ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 Article and publication are at http://www.proteinscience.org/cgi/doi/10.1110/ps.0213603. Reprint requests to: Knut Adermann, IPF PharmaCeuticals GmbH, Hannover, Germany; e-mail: knut.adermann@ipf-pharmaceuticals.de; fax: 49 (0) 511 5466 132.
ISSN:	0961-8368 1469-896X
DOI:	10.1110/ps.0213603