Chimeric Pesticidal Polypeptides for Insect Resistance
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Solution Overview
Problem
Current methods for controlling insect pests, such as corn rootworm, rely heavily on chemical pesticides, which pose environmental hazards and face challenges with resistance development, necessitating alternative solutions that enhance the efficacy of pesticidal polypeptides like Cry endotoxins from Bacillus spp.
Innovation Solution
The development of chimeric pesticidal polypeptides by fusing solubility-enhancing proteins with Cry endotoxins or their biologically active fragments, increasing their activity and stability, particularly in the gut environment of Coleopteran insects, and expressing these in plants to provide enhanced resistance against pests.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If traditional Cry endotoxins are used for pest control, then environmental hazards are reduced compared to chemical pesticides, but pest resistance develops and efficacy decreases
Solution Approach 1:
The patent modifies the amino acid sequence parameters of Cry endotoxins through site-directed mutagenesis at specific positions (e.g., positions 1-10, 11-20, 21-30, 31-40, 41-50, 51-60, 61-70, 71-80, 81-90, 91-100, 101-110, 111-120, 121-130, 131-140, 141-150, 151-160, 161-170, 171-180, 181-190, 191-200, 201-210, 211-220, 221-230, 231-240, 241-250, 251-260, 261-270, 271-280, 281-290, 291-300, 301-310, 311-320, 321-330, 331-340, 341-350, 351-360, 361-370, 371-380, 381-390, 391-400, 401-410, 411-420, 421-430, 431-440, 441-450, 451-460, 461-470, 471-480, 481-490, 491-500, 501-510, 511-520, 521-530, 531-540, 541-550, 551-560, 561-570, 571-580, 581-590, 591-600, 601-610, 611-620, 621-630, 631-640, 641-650, 651-660, 661-670, 671-680, 681-690, 691-700, 701-710, 711-720, 721-730, 731-740, 741-750, 751-760, 761-770, 771-780, 781-790, 791-800, 801-810, 811-820, 821-830, 831-840, 841-850, 851-860, 861-870, 871-880, 881-890, 891-900, 901-910, 911-920, 921-930, 931-940, 941-950, 951-960, 961-970, 971-980, 981-990, 991-1000) to create variants with enhanced pesticidal activity and reduced susceptibility to pest resistance. This parameter modification approach allows the endotoxins to maintain effectiveness against resistant pest populations.
Solution Approach 2:
The patent divides the Cry endotoxin protein into functional segments and regions, with specific focus on modifying particular amino acid sequences in defined portions of the protein structure. By segmenting the protein and targeting specific regions for mutagenesis, the invention can independently optimize different functional domains while maintaining overall protein stability and activity.
2Reliability
If site-directed mutagenesis is used to enhance Cry endotoxin activity, then pesticidal efficacy is improved, but the process becomes time-consuming and uncertain
Solution Approach 1:
The patent identifies and pre-determines specific amino acid positions that are most critical for pesticidal activity and pest resistance mechanisms. By focusing mutagenesis efforts on these pre-identified key positions rather than performing exhaustive random mutagenesis, the process time is significantly reduced while maintaining high probability of achieving desired efficacy improvements.
Solution Approach 2:
Instead of uniformly modifying the entire protein sequence, the patent applies localized mutagenesis to specific amino acid positions and regions that are known to be critical for function. This targeted approach concentrates evolutionary experimentation on the most promising areas, reducing the overall time and resources required while increasing the likelihood of successful outcomes.
3Reliability
If Cry endotoxins are expressed in plants to provide pest resistance, then crop protection is achieved, but the solubility and stability of the endotoxin in the plant gut environment is reduced
Solution Approach 1:
The patent introduces a solubility-enhancing polypeptide as an intermediary component that is co-expressed with the Cry endotoxin in the plant. This intermediary protein facilitates the solubility and stability of the endotoxin in the plant gut environment, acting as a molecular mediator that prevents aggregation and degradation while maintaining the endotoxin's pesticidal function.
Solution Approach 2:
The patent creates a composite protein system combining the Cry endotoxin with solubility-enhancing polypeptide sequences. This composite structure integrates the pesticidal functionality of the endotoxin with the solubility and stability properties of the enhancing polypeptide, resulting in a hybrid protein that maintains both protective and functional characteristics in the plant environment.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
The chimeric pesticidal polypeptides demonstrate improved activity against insect pests, including those resistant to traditional Cry endotoxins, offering increased solubility and efficacy when expressed in plants, thereby protecting crops from significant agricultural losses.
Implementation Method 1
The chimeric pesticidal polypeptides of the invention comprise a solubility-enhancing polypeptide fused to a Cry endotoxin or biologically active fragment thereof
Implementation Method 2
Once active, the endotoxins bind to the gut epithelium and form cation-selective channels that cause cell lysis and subsequent death
Implementation Method 3
form cation-selective channels that cause cell lysis and subsequent death
Data Source
AI summary
Methods and compositions for enhancing the resistance of plants to plant pests are provided. Chimeric pesticidal polypeptides and nucleic acid molecules encoding the chimeric pesticidal polypeptides are provided. The chimeric pesticidal polypeptides comprising a solubility-enhancing polypeptide operably linked to a polypeptide comprising pesticidal activity. The nucleic acid molecules can be used in expression cassettes for making transformed plants with enhanced resistance to plant pests. Further provided are transformed plants, plant tissues, plant cells, other host cells, and seeds as well as pesticidal compositions.