N-Glycosylation Profiles of the Green Microalgae Chlorella Zofingiensis

Abstract Nowadays, the use of microalgae species as raw materials in biopharmaceutical production is on the agenda. The reason behind this idea is that microalgae are cell factories that are able to efficiently utilize carbon dioxide for the production of numerous biologically active compounds. However, there are several problems that remain to be solved in the production of recombinant protein from microalgaes. One of the critical requirements is to produce a bio-compatible N-glycosylation profile from the secreted recombinant proteins. However, the knowledge about the glycosylation machinery and N-glycan profiles of microalgae spices are quite limited. In the study, it was aimed to characterize N-glycan profiles of a green microalgae, Chlorella zofingiensis. To achieve this, photoautotrophically grown Chlorella zofingiensis extracts including (glyco-)proteins were enzymatically deglycosylated and labelled with 2-aminobenzoic acid tag. Released N-glycans were purified with a HILIC-based approach and analyzed by MALDI-TOF(/TOF)-MS. The results showed that C. zofingiensis included oligomannosidic type N-glycan patterns. In addition, N-glycosylation profiles of C. zofingiensis by MALDI-MS revealed that most of the oligomannosidic N-glycans were phosphorylated. Oz Gunumuzde, mikroalg turlerinin biyofarmasotik uretiminde hammadde olarak kullanimi konusu tartisilmaktadir. .Bu fikrin altinda yatan temel neden mikro alglerin, biyolojik olarak aktif cok sayida bilesenin uretimi icin karbon dioksiti etkin bir sekilde kullanabilen hucre fabrikalari olmasidir. Bununla birlikte, mikroalglerden rekombinant protein uretiminde kullanilmasi icin cozulmesi gereken birkac sorun mevcuttur. Kritik gereksinimlerden birisi salgilanan rekombinant proteinlerden biyo-uyumlu bir N-glikozilasyon profile uretmektir. Ancak mikroalg turlerinin glikozilasyon makineleri ve N-glikan profilleri hakkindaki bilgiler oldukca sinirlidir. Bu calismada, yesil bir mikroalg turu olan Chlorella zofingiensis ‘in N-glikan profilinin karakterize edilmesi amaclandi. Bu amacla (gliko-)proteinleri iceren fotoototrofik olarak yetistirilmis Chlorella zofingiensis ekstreleri enzimatik olarak deglikozile edildi ve 2-aminobenzoik asit etiketi ile etiketlendi. Serbest hale getirilen N-glikanlar HILIC bazli bir yaklasimla saflastirildi ve MALDI-TOF (/TOF)-MS ile analiz edildi. Sonuclar C. zofingiensis'in oligomannozidik turde N-glikan modellerini icerdigini gostermistir. Ek olarak, MALDI-MS tarafindan C. zofingiensis'in N-glikosilasyon profilleri, oligomannosidik N-glikanlarin neredeyse yarisinin fosforilenmis oldugunu ortaya koymustur.