Recombinant Expression of a Novel Fungal Immunomodulatory Protein with Human Tumor Cell Antiproliferative Activity from Nectria haematococca

Abstract

To our best knowledge, all of the fungal immunomodulatory proteins (FIPs) have been successfully extracted and identified in Basidomycetes, with only the exception of FIP from ascomycete Nectria haematococca (FIP-nha) discovered through homology alignment most recently. In this work, a gene encoding FIP-nha was synthesized and recombinantly expressed in an Escherichia coli expression system. SDS-PAGE and MALDI-MS analyses of recombinant FIP-nha (rFIP-nha) indicated that the gene was successfully expressed. The yield of the bioactive FIP-nha protein was 42.7 mg/L. In vitro assays of biological activity indicated that the rFIP-nha caused hemagglutination of human and rabbit red blood cells, significantly stimulated mouse spleen lymphocyte proliferation, and enhanced expression of interleukin-2 (IL-2) released from mouse splenocytes, revealing a strong antitumor effect against HL60, HepG2 and MGC823. Through this work, we constructed a rapid and efficient method of FIP production, and suggested that FIP-nha is a valuable candidate for use in future medical care and pharmaceutical products.

Identification and functional characterization of a novel fungal immunomodulatory protein from Postia placenta

“…Characterization of the FIP-like sequence from the ascomycete Nectria haematococca was reported (Bastiaan-Net et al, 2013), while the manuscript was being prepared (Li et al, 2014). The other FIP-like sequence was found in the genome database of the basidiomycete P. placenta Mad-698-R (http://www.jgi.doe.gov/Postia).…”

Identification and Characterisation of a Novel Protein FIP-sch3 from Stachybotrys chartarum

“…placenta ) that were expressed in E . coli significantly inhibited the growth of several types of cancer cells and induced apoptosis, and the anti-tumour effects were cell-specific [18, 19]. These results indicated that, despite the sequence conservation between FIPs from different sources, their anti-tumour activity varied greatly.…”

“…Therefore, researchers have shifted away from the extraction or homologous cloning methods for identifying new FIPs and instead search for sequence similarity in the burgeoning genome databases. Two previously unknown FIP genes were quickly identified by genome mining and were largely produced using recombinant expression [17–19]. These two FIPs were very different from previously discovered FIPs: FIP-nha was the first FIP that was identified from the ascomycete Nectria haematococca , and FIP-ppl from Postia placenta was the only basidiomycete FIP to be identified outside the order of Macrofungi.…”

Fungal immunomodulatory proteins in the context of biomedicine

“…From that date, six other FIPs in species of the genus Ganoderma have been reported: FIP-gts of G. tsugae (15), FIPgsi of G. sinensis (16), FIP -gja of G. japonicum (Genbank: AAX9824), FIP-gmi G. Microsporum (17), FIP-gas of G. atrum (GenBank: KM077027.1) and FIP-gap G. applanatum (Genbank: AEP68179). Other eleven fungal immunomodulatory proteins have been described at molecular or sequence level in other Basidiomycetes: FIP-fve of Flammulina velutipes (18), FIP-vvo of straw mushroom (19), FIP-Apl of Auricularia polytricha (20), FIP-aca of Antrodia camphorate (21), FIP-PCIP Pleurotus citrinopileatus (22), FIP-PCP of Poria cocos (23), FIP-tvc of Trametes versicolor (24,25), FIP-ppl Postia placenta (26), FIPcru Chroogomphis rutilus (27), FIP-Lrh of Lignosus rhinocerotis (Tiger milk mushroom) (28), FIP-tfu of Tremella fuciformis (GenBank: EF152774) and FIPnha Nectria haemotococca edge Ascomycetes (29).…”

“…For several FIPs there is an experimental description of the basic DNA encoding sequence of the gene, and the primary, secondary and tertiary conformation of the protein (33,43,71). There are also insights about their effects on the immune system of vertebrate (29,69), however it is not known in depth the molecular mechanism that triggers the immunomodulatory response (26,72).…”