Biology subjects

Walter, G.

Publications and source records attributed to Walter, G..

3 recordsLinked to original sources

bioRxiv2025

Modular Nanobody Conjugates with Controlled Topology Using Genetically Encoded Non-canonical Amino Acids

Bispecific antibody-based derivatives are traditionally generated by fusing short, flexible peptides to two variable heavy and light chain pairs that recognize distinct antigens. However, this method limits domain joining to the amino (N) or carboxyl (C) termini, thereby restricting our ability to study the role of domain orientation in key properties of bispecific molecules, such as affinity and specificity. Here, we present an adaptable, plug-and-play application of genetic code expansion technology for the rapid, modular creation of bispecific nanobody conjugates from non-canonical amino acid (ncAA)- integrated nanobody domains, offering precise control over domain topology. We implement this strategy using a single-plasmid genetic code expansion system and computation-guided ncAA site selection to preserve nanobody expression and target binding. We demonstrate the effective incorporation and crosslinking of azide- and tetrazine-modified lysine and phenylalanine, respectively, at four engineered positions within an anti-PD-L1 nanobody and two positions within an anti-CTLA-4 nanobody in a panel of immune cell- and cancer-cell-binding nanobodies. Using this approach, we demonstrate the modular synthesis of a library of bispecific nanobodies, comprising four PD-L1xCD16 immune synapse engagers and eight PD-L1xCTLA-4 immune checkpoint bispecifics. This work enables a new dimension of control over protein crosslinking for rapidly constructing immune cell engagers for dual immune checkpoint blockade therapy and to bridge the synapse between immune cells and cancer cells.

bioengineering↗

bioRxiv2020

Tests of hybridisation in Tetragonula stingless bees using multiple genetic markers.

Discrepancies in mitochondrial and nuclear genetic data are often interpreted as evidence of hybridisation. We re-examined reports of hybridisation in three cryptic stingless bee species in the genus Tetragonula in South East Queensland, Australia (T. carbonaria, T. davenporti, and T. hockingsi). Previous studies on this group using microsatellite markers proposed that occasional hybrids are found. In contrast, we find that allele frequencies at neutral regions of the nuclear genome, both microsatellites and random snps, reliably separated the three species, and thus do not support hybridisation. We found no inter-species variation in PCR amplicons of the nuclear gene EF1alpha, but low and moderate species-specific polymorphisms in the nuclear gene Opsin and the mitochondrial 16S respectively, with no cases of mito-nuclear discordance at these genes. We confirm that nuclear divergence between these species is low, based on 10-26kb of non-coding sequence flanking EF1alpha and Opsin (0.7-1% pairwise difference between species). However, we find mitogenomes to be far more diverged than nuclear genomes (21.6-23.6% pairwise difference between species). Based on these comprehensive analyses of multiple marker types, we conclude that there is no ongoing gene flow in the Tetragonula species of South East Queensland, despite their high morphological similarity to one another and the low nuclear divergence among them. The mitogenomes and draft nuclear genomes provided for these species will be a resource for further molecular studies on this group, which are important pollinators in Australian natural and agroecosystems.

evolutionary biology↗

bioRxiv2020

Migration dynamics of an important rice pest: the brown planthopper (Nilaparvata lugens) across Asia - insights from population genomics

Brown planthoppers (Nilaparvata lugens) are the most serious insect pests of rice, one of the worlds most important staple crops. They reproduce year-round in the tropical parts of their distribution, but cannot overwinter in the temperate areas where they occur, and invade seasonally from elsewhere. Decades of research has not revealed their source unambiguously. We therefore sequenced the genomes of brown planthopper populations from across temperate and tropical parts of their distribution and show that the Indochinese peninsula is the major source of migration into temperate China. The Philippines, once considered a key source, is not significant, with little evidence for their migration into China. We find support for immigration from the west of China contributing to these regional dynamics. The lack of connectivity between the Philippines and mainland China explains the different evolution of Imidacloprid resistance in these populations. This study highlights the promise of whole genome sequence data to understand migration when gene flow is high - a situation that has been difficult to resolve using traditional genetic markers.

genomics↗