Structure determination of lipopeptides from Mycobacterium avium subspecies paratuberculosis and identification of antigenic lipopeptide probes.
Mycobacterium avium subspecies paratuberculosis (MAP) causes chronic illnesses mostly in ruminants. MAP infection of intestinal tissue triggers a fatal inflammatory disorder, Johne's disease (paratuberculosis). Development of fast and reliable diagnostic methods for Johne's disease in clinically suspected ruminants requires the discovery of MAP-specific antigens that induce immune responses. Despite a longtime interest in finding such antigens that can detect serum antibody responses with high sensitivity, the antigens currently used for a diagnosis of the MAP infections are the crude extracts from the whole cell. We performed the serum antibody response assay-guided purification of the ethanol extract from MAP isolated from an infected cow. With the results of extensive fractionations and in vitro assays, we identified that arachidyl-d-Phe-N-Me-l-Val-l-Ile-l-Phe-l-Ala-OH (named lipopeptide IIß, 3) exhibited the highest antibody binding activity in serum of a MAP-infected cattle compared with the other lipopeptides isolated from MAP. The absolute chemistry of 3 was determined unequivocally via our high-performance liquid chromatography (HPLC)-amino acid databases. α-Amino lipopeptide IIß and its fluorescent probes were synthesized and evaluated in serum antibody binding activity assays. Lipopeptide IIß-(2S)-NH2 (9) and its dansyl and fluorescein isothiocyanate (FITC) probes (10 and 11) exhibited antibody-mediated binding activity; thus, such MAP-specific lipopeptide probes can be potential biomarkers for the development of rapid and accurate diagnosis of Johne's disease.
Department of Pharmaceutical Sciences, College of Pharmacy, University of Tennessee Health Science Center, Memphis, TN, 38163, USA.
Katsuhiko
K
Mitachi
Department of Pharmaceutical Sciences, College of Pharmacy, University of Tennessee Health Science Center, Memphis, TN, 38163, USA.
Lekh Nath
LN
Sharma Gautam
Center for Wildlife Health, Department of Forestry, Wildlife, and Fisheries, University of Tennessee, Knoxville, TN, 37996, USA.
Jeffrey H
JH
Rice
Center for Wildlife Health, Department of Forestry, Wildlife, and Fisheries, University of Tennessee, Knoxville, TN, 37996, USA.
Keiko
K
Eda
Center for Wildlife Health, Department of Forestry, Wildlife, and Fisheries, University of Tennessee, Knoxville, TN, 37996, USA.
Ashutosh
A
Wadhwa
Department of Human Care, Tohto College of Health Sciences, Nishi Fukaya, Saitama, 366-0052, Japan.
Eiichi
E
Momotani
Center for Wildlife Health, Department of Forestry, Wildlife, and Fisheries, University of Tennessee, Knoxville, TN, 37996, USA.
Joseph P
JP
Hlopak
Center for Wildlife Health, Department of Forestry, Wildlife, and Fisheries, University of Tennessee, Knoxville, TN, 37996, USA.
Shigetoshi
S
Eda
Department of Pharmaceutical Sciences, College of Pharmacy, University of Tennessee Health Science Center, Memphis, TN, 38163, USA. Electronic address: mkurosu@uthsc.edu.
Michio
M
Kurosu
D000818:Animals; D000907:Antibodies, Bacterial / Q000737:chemistry / Q000276:immunology; D000942:Antigens, Bacterial / Q000737:chemistry* / Q000276:immunology*; D002417:Cattle; D005456:Fluorescent Dyes / Q000138:chemical synthesis / Q000737:chemistry*; D055666:Lipopeptides / Q000737:chemistry* / Q000276:immunology*; D016927:Mycobacterium avium subsp. paratuberculosis / Q000737:chemistry* / Q000276:immunology; D011487:Protein Conformation
D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D005456:Fluorescent Dyes; D055666:Lipopeptides
Johne's disease; Lipopeptide; Lipopeptide probes; Mycobacterium avium subspecies paratuberculosis; New chiral reprivatizing agents