In-silico determination of Pichia pastoris signal peptides for extracellular recombinant protein production

Massahi, Aslan; Çalı́k, Pınar

doi:10.1016/j.jtbi.2014.08.048

Cited by 30 publications

(9 citation statements)

References 52 publications

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“…Signal peptide (SP) design and its synthetic construction is an effective method used in industrial biotechnology to improve heterologous protein production and biocatalysis. 72 , 73 Thus, investigation of the differential codon composition of genes with SPs has been an objective of various studies despite the different approaches employed. 26 , 43–44 We used our integrated framework, to analyse the CDSs containing SPs for each yeast genome, by computing the Y-value ( Fig.…”

Section: Resultsmentioning

confidence: 99%

Integrated analysis of individual codon contribution to protein biosynthesis reveals a new approach to improving the basis of rational gene design

2017

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Gene codon optimization may be impaired by the misinterpretation of frequency and optimality of codons. Although recent studies have revealed the effects of codon usage bias (CUB) on protein biosynthesis, an integrated perspective of the biological role of individual codons remains unknown. Unlike other previous studies, we show, through an integrated framework that attributes of codons such as frequency, optimality and positional dependency should be combined to unveil individual codon contribution for protein biosynthesis. We designed a codon quantification method for assessing CUB as a function of position within genes with a novel constraint: the relativity of position-dependent codon usage shaped by coding sequence length. Thus, we propose a new way of identifying the enrichment, depletion and non-uniform positional distribution of codons in different regions of yeast genes. We clustered codons that shared attributes of frequency and optimality. The cluster of non-optimal codons with rare occurrence displayed two remarkable characteristics: higher codon decoding time than frequent–non-optimal cluster and enrichment at the 5′-end region, where optimal codons with the highest frequency are depleted. Interestingly, frequent codons with non-optimal adaptation to tRNAs are uniformly distributed in the Saccharomyces cerevisiae genes, suggesting their determinant role as a speed regulator in protein elongation.

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Section: Resultsmentioning

confidence: 99%

Integrated analysis of individual codon contribution to protein biosynthesis reveals a new approach to improving the basis of rational gene design

2017

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“…SP1 and SP2 therefore were selected as the top two emitted peptides for proceeding onto further structural analyses. Prior to structural analyses, we also examined the sequences for putative N-glycosylation sites using NetNGlyc1.0, as previously described [24]. Our analyses showed an absence of glycosylation sites in both the emitted SP and native SP, which would be important to avoid interference with Signal Peptidase I processing events.…”

Section: Resultsmentioning

confidence: 99%

Skeletal muscle signal peptide optimization for enhancing propeptide or cytokine secretion

Neto

Figueiredo

2016

Journal of Theoretical Biology

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We have utilized hidden Markov models using HMMER software to predict and generate putative strong secretory signal peptide sequences for directing efficient secretion of cytokines from skeletal muscle for therapeutic applications. The results show that this approach can analyze signal sequences of a skeletal muscle secretome dataset and classify them, emitting new sequences that are strong candidate skeletal muscle-enriched signal peptides. The emitted signal peptides also were analyzed for their hydropathy and secondary structure profiles as compared to native signal peptides. The emitted signal peptides had a higher degree of hydropathy and helical composition relative to native sequences, which may suggest that these new sequences may hold promise for promoting enhanced secretion of proteins including cytokines or propeptides from skeletal muscle.

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“…23 This has been attributed to bottlenecks in the secretory pathway which may become overwhelmed by the effects of high titers of a particular heterologous protein. 24 Strategies to overcome these barriers may include the use of new signal peptides based on the K. phaffii secretome 25 and combinatorial engineering of secretion helper factors involved in protein folding and vesicle trafficking. 26 In K. phaffii, expression vectors are typically integrated into the host chromosome by homologous recombination and non-homologous end joining events.…”

Section: Multi-copy Number Clonesmentioning

confidence: 99%

Molecular strategies to increase the levels of heterologous transcripts inKomagataella phaffiifor protein production

et al. 2017

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Komagataella phaffii (formerly Pichia pastoris) is a well-known fungal system for heterologous protein production in the context of modern biotechnology. To obtain higher protein titers in this system many researchers have sought to optimize gene expression by increasing the levels of transcription of the heterologous gene. This has been typically achieved by manipulating promoter sequences or by generating clones bearing multiple copies of the desired gene. The aim of this work is to describe how these different molecular strategies have been applied in K. phaffii presenting their advantages and drawbacks.

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In-silico determination of Pichia pastoris signal peptides for extracellular recombinant protein production

Cited by 30 publications

References 52 publications

Integrated analysis of individual codon contribution to protein biosynthesis reveals a new approach to improving the basis of rational gene design

Integrated analysis of individual codon contribution to protein biosynthesis reveals a new approach to improving the basis of rational gene design

Skeletal muscle signal peptide optimization for enhancing propeptide or cytokine secretion

Molecular strategies to increase the levels of heterologous transcripts inKomagataella phaffiifor protein production

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