Extracellular expression of YlLip11 with a native signal peptide from Yarrowia lipolytica MSR80 in three different yeast hosts.
Lipase YlLip11 from Yarrowia lipolytica was expressed with a signal peptide encoding sequence in Arxula adeninivorans, Saccharomyces cerevisiae and Hansenula polymorpha using the Xplor®2 transformation/expression platform and an expression module with the constitutive Arxula-derived TEF1 promoter. The YlLip11 signal peptide was functional in all of the yeast hosts with 97% of the recombinant enzyme being secreted into the culture medium. However, recombinant YlLip11 with His Tag fused at C-terminal was not active. The best recombinant YlLip11 producing A. adeninivorans G1212/YRC102-YlLip11 transformant cultivated in shake flasks produced 2654 U/L lipase, followed by S. cerevisiae SEY6210/YRC103-YlLip11 (1632U/L) and H. polymorpha RB11/YRC103-YlLip11 (1144U/L). Although the biochemical parameters of YlLip11 synthesized in different hosts were similar, their glycosylation level and thermo stability differed. The protein synthesized by the H. polymorpha transformant had the highest degree of glycosylation and with a t1/2 of 60min at 70°C, exhibited the highest thermostability.
Department of Microbiology, University of Delhi South Campus, New Delhi 110021, India.
Arti
A
Kumari
School of Biological Sciences, University of Canterbury, Private Bag 4800, Christchurch, New Zealand.
Keith
K
Baronian
Leibniz-Institut für Pflanzengenetik und Kulturpflanzenforschung (IPK), Corrensstr. 3, 06466 Gatersleben, Germany. Electronic address: kunzeg@ipk-gatersleben.de.
Gotthard
G
Kunze
Department of Microbiology, University of Delhi South Campus, New Delhi 110021, India. Electronic address: ranigupta15@rediffmail.com.
Rani
R
Gupta
D003001:Cloning, Molecular; D004795:Enzyme Stability; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D015870:Gene Expression; D006031:Glycosylation; D006639:Histidine / Q000737:chemistry / Q000235:genetics; D008049:Lipase / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D009842:Oligopeptides / Q000737:chemistry / Q000235:genetics; D010843:Pichia / Q000737:chemistry / Q000201:enzymology / Q000235:genetics*; D010957:Plasmids / Q000737:chemistry / Q000378:metabolism; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000737:chemistry / Q000201:enzymology / Q000235:genetics*; D004718:Saccharomycetales / Q000737:chemistry / Q000201:enzymology / Q000235:genetics*; D013045:Species Specificity; D014170:Transformation, Genetic; D025062:Yarrowia / Q000737:chemistry / Q000201:enzymology / Q000235:genetics*
D005656:Fungal Proteins; C471213:His-His-His-His-His-His; D009842:Oligopeptides; D021382:Protein Sorting Signals; D011993:Recombinant Fusion Proteins; D006639:Histidine; D008049:Lipase
Arxula adeninivorans; Hansenula polymorpha; Lipase; Saccharomyces cerevisiae; Yarrowia lipolytica