The Purification Of Antifungal Substances And Genome Cluster Analysis Of Paenibacillus Polymyxa KF-1

The strain KF-1 was isolated from our laboratory, and it was identified as a strain of Paenibacillus polymyxa through physiological and biochemical characteristics and 16 S rDNA sequence, KF-1 exhibited the ability of inhibiting various fungus.. The antimicrobial active substance secreted by KF-1 was extracted using ethyl acetate, then purified by high performance liquid chromatograph. Through electrospray ionization mass spectrometry(ESI-MS) analysis, four groups of antimicrobial active substances were found: the Peak 1 with a retention time of 14.753 min, and a molecular weight of 188 was probably methyl 3,5-dihydroxybenzoate; Peak 2 exhibited a retention time of 22.676 min, and the yield was very high. For gram-positive bacteria and fungi, this compound exhibited obvious inhibition. The ion peak [M+Na]+ m/z and the double peak [2M+Na]+ m/z of this compound were 413.8 and 803.6 respectively. Combined with a molecular weight of 390, we guess it was a large ring esters antibiotics with small molecule of; Peak 3 includes two sets of compounds, of which molecular weights were 251 and 453 seperately. Compared with known antibiotics, it was similar with the nucleoside antibiotics Polyoxin. According to their abilities of inhibiting normal growth of candida albicans, it may belong to antifungal antibiotics. Peak 4 mainly includes two compounds with molecular weights of 882.4 and 933.5, which belong to LI-F lipopeptide family Fusaridins A [M+H]+ LI-07 F respectively.The genome sequence of Paenibacillus sp KF-1 was then analyzed by whole-genome shotgun method. The full length of KF-1 genome was 945960471 bp, and the GC content was 46.26%. There were 5229 ORF in the genome of KF-1, which represents 84.53% of the total length of genome.KF-1 contains six groups of 16S/5S/23 S rRNA operon regions, and 107 non-coding tRNA was also identified,with a total length of 8275 bp. The sequence information of KF-1 has been submitted to GenBank, and the accession number is LNZF00000000. By GO and eggnog annotations,many ORF in the KF-1 genome encoding transport proteins involved in the metabolism of amino acids, transport of nucleotides, and synthesis of antibiotics. 22 different gene clusters were predicted to be involved in the synthesis of in KF-with the method of antiSMASH. and. Some of these secondary metabolites showed high homology to Iturin, Fusaricidin, colistin, trdecaptin, and polymyxin antibiotics, while some of them was not clear. These compounds with antibacterial activity, were mainly synthesized by non-ribosomal peptide synthase(NPRS), non-ribosomal peptide synthase combined with polypeptide synthase(PKS), and Lantipeptide pathway respectively.Therefore, Paenibacillus KF-1 is a beneficial bacteria which can produce various antifungal active substancesand it has a very broad application prospect.