Biology subjects

Greaves, J.

Publications and source records attributed to Greaves, J..

6 recordsLinked to original sources

bioRxiv2026

AI-derived Protein Structures Validation: AlphaFold2 Models in the Twilight Zone

In any field, unquestioningly accepting artificial intelligence (AI) results should be considered bad practise. Here, we devised a comparative modelling-based strategy for validating protein structures that exploits the well-known observation that protein folds are far more conserved than protein sequences. We identify proteins with a similar fold to the AlphaFold-generated query protein and determine their structural alignment to the query. The hypothesis is that if the sequence alignment coincides with the structural alignment, then the structure is validated. The strategy is implemented on a helix-by-helix and strand-by-strand basis using a multi-template pairwise local profile alignment method that works well into the twilight zone. The method is illustrated by application to the transmembrane transporter PEPT1, for which the structure is known, and the S-deacylases ABHD13 and ABHD16A, for which only AI-generated models exist. ABHD16A is particularly challenging because a sequence alignment search with BLASTp does not reveal any structural homologues and therefore requires work with extremely remote homologues; however, both models are validated through this strategy and are stable during classical molecular dynamics simulations. The ability of the strategy to identify errors is assessed with reference to misaligned ABHD13 models and misfolded decoy proteins.

bioinformatics↗

bioRxiv2026

Deacylation of SNAP25 protein family isoforms reveals distinct substrate selectivities of α/β hydrolase domain (ABHD) deacylases

Reversible S-acylation controls protein trafficking, localisation and activity within cells, yet the limited understanding of the regulatory mechanisms of S-acylation/deacylation cycles represents one of the biggest obstacles in exploiting this prevalent post-translational modification for therapeutic purposes. Here, we identified ABHD13 and ABHD16A as novel deacylases regulating the SNAP25 family of SNARE proteins responsible for vesicular membrane fusion. We determined the mechanism and structural determinants for ABHD13-mediated deacylation and substrate selectivity towards SNAP25b, and identified a novel mechanism of acyl chain extraction for integral membrane deacylases. This is the first report of deacylase selectivity mapped to a single amino acid position in full-length proteins.

molecular biology↗

bioRxiv2022

Comparable human spatial memory distortions in physical, desktop virtual and immersive virtual environments

Boundaries define regions of space and are integral to episodic memories. The impact of boundaries on spatial memory and neural representations of space has been extensively studied in freely-moving rodents. But less is known in humans and many prior studies have employed desktop virtual reality (VR) which lacks the body-based self-motion cues of the physical world, diminishing the potentially strong input from path integration to spatial memory. We replicated a desktop-VR study testing the impact of boundaries on spatial memory (Hartley et al., 2004) in a physical room (2.4m x 2.4m, 2m tall) by having participants (N = 27) learn the location of a circular stool and then after a short delay replace it where they thought they had found it. During the delay, the wall boundaries were either expanded or contracted. We compared performance to groups of participants undergoing the same procedure in a laser-scanned replica in both desktop VR (N = 44) and freely-walking head mounted display (HMD) VR (N = 39) environments. Performance was measured as goodness of fit between the spatial distributions of group responses and seven modelled distributions that prioritised different metrics based on boundary geometry or walking paths to estimate the stool location. The best fitting model was a weighted linear combination of all the geometric spatial models, but an individual model derived from place cell firing in Hartley et al. 2004 also fit well. High levels of disorientation in all three environments prevented detailed analysis on the contribution of path integration. We found identical model fits across the three environments, though desktop VR and HMD-VR appeared more consistent in spatial distributions of group responses than the physical environment and displayed known variations in virtual depth perception. Thus, while human spatial representation appears differentially influenced by environmental boundaries, the influence is similar across virtual and physical environments. Despite differences in body-based cue availability, desktop and HMD-VR allow a good and interchangeable approximation for examining human spatial memory in small-scale physical environments.

neuroscience↗

bioRxiv2021

Viability of MS2 and Phi6 Bacteriophages on Carpet and Dust

Respiratory viral illnesses are commonly spread in the indoor environment through multiple transmission routes, including droplets, aerosols, and direct/indirect contact. Indoors, resuspension of dust from flooring is a major source of human exposure. However, it is critical to determine viral persistence on dust and flooring to better characterize human exposure. The goal of this work is to determine viral viability on two carpet types (cut and looped) and house dust over time and after four different cleaning methods. MS2 and Phi6 bacteriophages were used to represent non-enveloped and enveloped viruses, respectively. These viral surrogates were placed in an artificial saliva solution and nebulized onto carpet or dust. Viability was measured at various time points (0, 1, 2, 3, 4, 24, and 48 hours) and after cleaning (vacuuming, hot water extraction with stain remover, steam, and a disinfection spray). Viability decay was modeled as first-order. MS2 bacteriophages showed slower viability decay rates in dust (-0.11 hr-1), cut carpet (-0.20 hr-1), and looped carpet (-0.09 hr-1) compared to Phi6 (-3.36 hr-1, -1.57 hr-1, and - 0.20 hr-1 respectively). The difference between phages was statistically significant in dust and cut carpet (p<0.05). Viral RNA demonstrated minimal degradation that in most cases was not statistically different from zero over the 48 hours measured (p>0.05). Viable viral concentrations were reduced to below the detection limit for steam and disinfection for both MS2 and Phi6 (p<0.05), while vacuuming and hot water extraction with stain remover showed no significant changes in concentration from uncleaned carpet (p>0.05). This study used viral surrogates and did not model risk of viral transmission via dust. Overall, these results demonstrate that MS2 and Phi6 bacteriophages can remain viable in carpet and dust for several hours to days, and cleaning techniques with heat and disinfectants may be more effective than standard vacuuming for viral removal. Future work should model risk from exposure via dust and flooring for various viruses such as influenza, SARS-CoV-2, and RSV. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=99 SRC="FIGDIR/small/444479v1_ufig1.gif" ALT="Figure 1"> View larger version (31K): org.highwire.dtl.DTLVardef@1cc0ad9org.highwire.dtl.DTLVardef@dc440corg.highwire.dtl.DTLVardef@f787dcorg.highwire.dtl.DTLVardef@f8bda3_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2020

The Linker Domain of SNAP25 Acts as a Flexible Molecular Spacer to Ensure Efficient S-Acylation

S-Acylation of the SNARE protein SNAP25 is mediated by a subset of Golgi zDHHC enzymes, in particular zDHHC17. The ankyrin repeat (ANK) domain of this enzyme interacts with a short linear motif known as the zDHHC ANK binding motif (zDABM) in SNAP25 (112-VVASQP-117), which is downstream of the S-acylated cysteine-rich domain (85-CGLCVCPC-92). In this study, we have investigated the importance of the flexible linker (amino acids 93-111; referred to as the "mini-linker" region) that separates the zDABM and S-acylated cysteines. Shortening the mini-linker had no effect of zDHHC17 interaction but blocked S-acylation. Insertion of additional flexible glycine-serine repeats had no effect on S-acylation, whereas extended and rigid alanine-proline repeats perturbed this process. Indeed, a SNAP25 mutant in which the mini-linker region was substituted with a flexible glycine-serine linker of the same length underwent efficient S-acylation. Furthermore, this mutant displayed the same intracellular localisation as wild-type SNAP25, showing that the sequence of the mini-linker is not important in this context. By using the results of previous peptide array experiments, we generated a SNAP25 mutant predicted to have a higher affinity zDABM, and this mutant showed enhanced interaction with zDHHC17 in cells. Interestingly, this mutant was S-acylated with reduced efficiency, implying that a lower affinity interaction of the SNAP25 zDABM with zDHHC17 is optimal for S-acylation efficiency. Overall, the results of this study show that amino acids 93-111 in SNAP25 act as a flexible molecular spacer to ensure efficient coupling of enzyme-substrate interaction and S-acylation.

cell biology↗

bioRxiv2017

CYRI (FAM49) proteins are local inhibitors of Scar/WAVE induced lamellipodia that bind directly to active Rac1

Actin-based protrusions driving cell migration are reinforced through positive feedback, but it is unclear how the cell restricts the eventual size of a protrusion or limits positive signals to cause splitting or retraction. We have identified an evolutionarily conserved regulator of the protrusion machinery, which we name CYRI (CYFIP-related Rac interacting) protein. CYRI shows sequence similarity to the Scar/WAVE complex member CYFIP in a Domain of Unknown Function, DUF1394. CYRI binds specifically to activated Rac1 via a common motif shared with CYFIP, establishing DUF1394 as a new Rac1 binding domain. CYRI-depleted cells have broad, Scar/WAVE-enriched lamellipodia and enhanced Rac1 signaling. Conversely, CYRI overexpression suppresses spreading and dramatically sharpens protrusions into unproductive needles. CYRI proteins use dynamic inhibition of Scar/WAVE induced actin to focus positive protrusion signals and regulate pseudopod complexity. CYRI behaves like a \"local inhibitor\" predicted and described in widely accepted mathematical models, but not previously identified in living cells.

cell biology↗