GASP: A Pan-Specific Predictor of Family 1 Glycosyltransferase Acceptor Specificity Enabled by a Pipeline for Substrate Feature Generation and Large-Scale Experimental Screening.

Glycosylation represents a major chemical challenge; while it is one of the most common reactions in Nature, conventional chemistry struggles with stereochemistry, regioselectivity, and solubility issues. In contrast, family 1 glycosyltransferase (GT1) enzymes can glycosylate virtually any given nucleophilic group with perfect control over stereochemistry and regioselectivity. However, the appropriate catalyst for a given reaction needs to be identified among the tens of thousands of available sequences. Here, we present the glycosyltransferase acceptor specificity predictor (GASP) model, a data-driven approach to the identification of reactive GT1:acceptor pairs. We trained a random forest-based acceptor predictor on literature data and validated it on independent in-house generated data on 1001 GT1:acceptor pairs, obtaining an AUROC of 0.79 and a balanced accuracy of 72%. The performance was stable even in the case of completely new GT1s and acceptors not present in the training data set, highlighting the pan-specificity of GASP. Moreover, the model is capable of parsing all known GT1 sequences, as well as all chemicals, the latter through a pipeline for the generation of 153 chemical features for a given molecule taking the CID or SMILES as input (freely available at https://github.com/degnbol/GASP). To investigate the power of GASP, the model prediction probability scores were compared to GT1 substrate conversion yields from a newly published data set, with the top 50% of GASP predictions corresponding to reactions with >50% synthetic yields. The model was also tested in two comparative case studies: glycosylation of the antihelminth drug niclosamide and the plant defensive compound DIBOA. In the first study, the model achieved an 83% hit rate, outperforming a hit rate of 53% from a random selection assay. In the second case study, the hit rate of GASP was 50%, and while being lower than the hit rate of 83% using expert-selected enzymes, it provides a reasonable performance for the cases when an expert opinion is unavailable. The hierarchal importance of the generated chemical features was investigated by negative feature selection, revealing properties related to cyclization and atom hybridization status to be the most important characteristics for accurate prediction. Our study provides a GT1:acceptor predictor which can be trained on other data sets enabled by the automated feature generation pipelines. We also release the new in-house generated data set used for testing of GASP to facilitate the future development of GT1 activity predictors and their robust benchmarking.

Systematic inference of indirect transcriptional regulation by protein kinases and phosphatases.

Gene expression is controlled by pathways of regulatory factors often involving the activity of protein kinases on transcription factor proteins. Despite this well established mechanism, the number of well described pathways that include the regulatory role of protein kinases on transcription factors is surprisingly scarce in eukaryotes. To address this, PhosTF was developed to infer functional regulatory interactions and pathways in both simulated and real biological networks, based on linear cyclic causal models with latent variables. GeneNetWeaverPhos, an extension of GeneNetWeaver, was developed to allow the simulation of perturbations in known networks that included the activity of protein kinases and phosphatases on gene regulation. Over 2000 genome-wide gene expression profiles, where the loss or gain of regulatory genes could be observed to perturb gene regulation, were then used to infer the existence of regulatory interactions, and their mode of regulation in the budding yeast Saccharomyces cerevisiae. Despite the additional complexity, our inference performed comparably to the best methods that inferred transcription factor regulation assessed in the DREAM4 challenge on similar simulated networks. Inference on integrated genome-scale data sets for yeast identified ∼ 8800 protein kinase/phosphatase-transcription factor interactions and ∼ 6500 interactions among protein kinases and/or phosphatases. Both types of regulatory predictions captured statistically significant numbers of known interactions of their type. Surprisingly, kinases and phosphatases regulated transcription factors by a negative mode or regulation (deactivation) in over 70% of the predictions.