Interleukin-2 superkines by computational design

Ren, Junming; Chu, Alexander E.; Jude, Kevin; Picton, Lora K.; Kare, Aris J; Su, Leon; Romero, Alejandra Montano; Huang, Po-Ssu; García, K. Christopher

doi:10.1073/pnas.2117401119

Cited by 15 publications

(8 citation statements)

References 47 publications

(53 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…2 and Box 1). Although these computational-based cytokine redesigns target the cytokine-receptor interface, new affinity-matured cytokines can also be engineered solely through computational-driven protein stabilization, as showcased in a proof-of-concept study with a new IL-2Rβ-selective variant 114 . This computational protein stabilization approach requires less experimental intervention compared to the receptor-interface-focused designs, such as STK-012 and NL-201, potentially decreasing the costs and time involved in computational cytokine engineering.…”

Section: Review Articlementioning

confidence: 99%

Engineering cytokine therapeutics

Deckers¹,

Anbergen²,

Hokke

et al. 2023

Nat Rev Bioeng

View full text Add to dashboard Cite

Cytokines have pivotal roles in immunity, making them attractive as therapeutics for a variety of immune-related disorders. However, the widespread clinical use of cytokines has been limited by their short blood half-lives and severe side effects caused by low specificity and unfavourable biodistribution. Innovations in bioengineering have aided in advancing our knowledge of cytokine biology and yielded new technologies for cytokine engineering. In this Review, we discuss how the development of bioanalytical methods, such as sequencing and high-resolution imaging combined with genetic techniques, have facilitated a better understanding of cytokine biology. We then present an overview of therapeutics arising from cytokine re-engineering, targeting and delivery, mRNA therapeutics and cell therapy. We also highlight the application of these strategies to adjust the immunological imbalance in different immune-mediated disorders, including cancer, infection and autoimmune diseases. Finally, we look ahead to the hurdles that must be overcome before cytokine therapeutics can live up to their full potential.

show abstract

Section: Review Articlementioning

confidence: 99%

Engineering cytokine therapeutics

Deckers¹,

Anbergen²,

Hokke

et al. 2023

Nat Rev Bioeng

View full text Add to dashboard Cite

show abstract

“…Sequences were designed in Rosetta using an iterative enrichment procedure in which the Monte Carlo search space was narrowed down in successive rounds of design, based on amino acids enriched, or more frequently selected by the design algorithm, at each position [ 15 , 26 ]. During the sequence design process, we observed many different barrel eccentricities, suggesting that our syntax is amenable to more curvatures than the ones designed here (see Materials and Methods).…”

Section: Resultsmentioning

confidence: 99%

“…First, automated design was conducted with the relax/minimization step constrained to starting coordinates, to prevent changes to the β barrel curvature. Once the backbone architecture was stabilized, we refined the sequence using three rounds of an “iterative enrichment” protocol [ 26 ]. Each residue position was classified by manual inspection as a hydrophobic, solvent-exposed, or boundary (could be hydrophobic or polar) position.…”

Section: Methodsmentioning

confidence: 99%

De Novo Design of a Highly Stable Ovoid TIM Barrel: Unlocking Pocket Shape towards Functional Design

et al. 2022

Self Cite

View full text Add to dashboard Cite

The ability to finely control the structure of protein folds is an important prerequisite to functional protein design. The TIM barrel fold is an important target for these efforts as it is highly enriched for diverse functions in nature. Although a TIM barrel protein has been designed de novo, the ability to finely alter the curvature of the central beta barrel and the overall architecture of the fold remains elusive, limiting its utility for functional design. Here, we report the de novo design of a TIM barrel with ovoid (twofold) symmetry, drawing inspiration from natural beta and TIM barrels with ovoid curvature. We use an autoregressive backbone sampling strategy to implement our hypothesis for elongated barrel curvature, followed by an iterative enrichment sequence design protocol to obtain sequences which yield a high proportion of successfully folding designs. Designed sequences are highly stable and fold to the designed barrel curvature as determined by a 2.1 Å resolution crystal structure. The designs show robustness to drastic mutations, retaining high melting temperatures even when multiple charged residues are buried in the hydrophobic core or when the hydrophobic core is ablated to alanine. As a scaffold with a greater capacity for hosting diverse hydrogen bonding networks and installation of binding pockets or active sites, the ovoid TIM barrel represents a major step towards the de novo design of functional TIM barrels.

show abstract

“…Protein engineering approaches (experimental and computational) focused on structure-guided stabilization of metastable regions, rather than on the receptor binding interfaces, have been successfully applied to affinity maturation studies on IL-2. 82,83 Thermostable "IL-2 superkine" variants with increased affinity for the IL-2Rβ receptor were produced either by stabilizing protein loops, improving protein packing and/or redesigning the hydrophobic core of the protein.…”

Section: Molecular Dynamics Simulationsmentioning

confidence: 99%

Lessons on protein structure from interleukin‐4: All disulfides are not created equal

Vaz,

Rodrigues,

Loureiro‐Ferreira

et al. 2023

Proteins

View full text Add to dashboard Cite

Interleukin‐4 (IL‐4) is a hematopoietic cytokine composed by a four‐helix bundle stabilized by an antiparallel beta‐sheet and three disulfide bonds: Cys3‐Cys127, Cys24‐Cys65, and Cys46‐Cys99. IL‐4 is involved in several immune responses associated to infection, allergy, autoimmunity, and cancer. Besides its physiological relevance, IL‐4 is often used as a “model” for protein design and engineering. Hence, to understand the role of each disulfide in the structure and dynamics of IL‐4, we carried out several spectroscopic analyses (circular dichroism [CD], fluorescence, nuclear magnetic resonance [NMR]), and molecular dynamics (MD) simulations on wild‐type IL‐4 and four IL‐4 disulfide mutants. All disulfide mutants showed loss of structure, altered interhelical angles, and looser core packings, showing that all disulfides are relevant for maintaining the overall fold and stability of the four‐helix bundle motif, even at very low pH. In the absence of the disulfide connecting both protein termini Cys3‐Cys127, C3T‐IL4 showed a less packed protein core, loss of secondary structure (~9%) and fast motions on the sub‐nanosecond time scale (lower S2 order parameters and larger τc correlation time), especially at the two protein termini, loops, beginning of helix A and end of helix D. In the absence of Cys24‐Cys65, C24T‐IL4 presented shorter alpha‐helices (14% loss in helical content), altered interhelical angles, less propensity to form the small anti‐parallel beta‐sheet and increased dynamics. Simultaneously deprived of two disulfides (Cys3‐Cys127 and Cys24‐Cys65), IL‐4 formed a partially folded “molten globule” with high 8‐anilino‐1‐naphtalenesulphonic acid‐binding affinity and considerable loss of secondary structure (~50%decrease), as shown by the far UV‐CD, NMR, and MD data.

show abstract

Interleukin-2 superkines by computational design

Cited by 15 publications

References 47 publications

Engineering cytokine therapeutics

Engineering cytokine therapeutics

De Novo Design of a Highly Stable Ovoid TIM Barrel: Unlocking Pocket Shape towards Functional Design

Lessons on protein structure from interleukin‐4: All disulfides are not created equal

Contact Info

Product

Resources

About