An Educational Bioinformatics Project to Improve Genome Annotation

Abstract

Scientific advancement is hindered without proper genome annotation because biologists lack a complete understanding of cellular protein functions. In bacterial cells, hypothetical proteins (HPs) are open reading frames with unknown functions. HPs result from either an outdated database or insufficient experimental evidence (i.e., indeterminate annotation). While automated annotation reviews help keep genome annotation up to date, often manual reviews are needed to verify proper annotation. Students can provide the manual review necessary to improve genome annotation. This paper outlines an innovative classroom project that determines if HPs have outdated or indeterminate annotation. The Hypothetical Protein Characterization Project uses multiple well-documented, freely available, web-based, bioinformatics resources that analyze an amino acid sequence to (1) detect sequence similarities to other proteins, (2) identify domains, (3) predict tertiary structure including active site characterization and potential binding ligands, and (4) determine cellular location. Enough evidence can be generated from these analyses to support re-annotation of HPs or prioritize HPs for experimental examinations such as structural determination via X-ray crystallography. Additionally, this paper details several approaches for selecting HPs to characterize using the Hypothetical Protein Characterization Project. These approaches include student- and instructor-directed random selection, selection using differential gene expression from mRNA expression data, and selection based on phylogenetic relations. This paper also provides additional resources to support instructional use of the Hypothetical Protein Characterization Project, such as example assignment instructions with grading rubrics, links to training videos in YouTube, and several step-by-step example projects to demonstrate and interpret the range of achievable results that students might encounter. Educational use of the Hypothetical Protein Characterization Project provides students with an opportunity to learn and apply knowledge of bioinformatic programs to address scientific questions. The project is highly customizable in that HP selection and analysis can be specifically formulated based on the scope and purpose of each student’s investigations. Programs used for HP analysis can be easily adapted to course learning objectives. The project can be used in both online and in-seat instruction for a wide variety of undergraduate and graduate classes as well as undergraduate capstone, honor’s, and experiential learning projects.

Genome-Wide Analysis of miR159 Gene Family and Predicted Target Genes Associated with Environmental Stress in Dendrobium officinale: A Bioinformatics Study

“…These findings suggested that GAMYB genes could be differently regulated under different stresses, which might result from the expression change of corresponding miRNA159s. Most proteins can be functionally annotated via an experimental and computational approach, but some proteins with unknown domains are referred to as hypothetical proteins [ 46 ]. Dca019932 , one of the predicted Dof-miR159-targeted genes encoding the hypothetical protein, was enriched in terms of DNA-binding transcription factor activity by the GO analysis ( Table S3 ).…”

TargetingLeishmania infantumMannosyl-oligosaccharide glucosidase with natural products: pH-dependent inhibition explored through computer-aided drug design

“…Notably, in early genome sequencing endeavors, numerous computationally predicted open reading frames (ORFs) encoded proteins lacking functional annotation, called hypothetical proteins (Amatore et al, 2020). An ORF that exhibited sequence homology to enzymes in the Glycoside Hydrolase (GH) family attracted attention.…”

“…Due to the promising results obtained in our previous research (Chavez-Fumagalli et al, 2019; Costa et al, 2021), we utilized and modeled MOGS (formerly categorized as a hypothetical protein) as a potential target and receptor in this study to evaluate the effect of NPs (Figure 3A). Notably, in early genome sequencing endeavors, numerous computationally predicted open reading frames (ORFs) encoded proteins lacking functional annotation, called hypothetical proteins (Amatore et al, 2020). An ORF that exhibited sequence homology to enzymes in the Glycoside Hydrolase (GH) family attracted attention.…”

Antifungal and Aflatoxin-Reducing Activity of β-Glucan Isolated from Pichia norvegensis Grown on Tofu Wastewater

“…The FASTA contig data were then aligned using the Basic Local Alignment Search Tool (BLAST) on the NCBI website to compare the nucleotide sequences in Genbank and calculate their statistical match so that the yeast species with the highest identification could be identified. The characteristics of sequences that had high identification had the same maximum score and total score, query coverage up to 100%, e-value of 0.0 (zero), and percentage identify of 100% [51].…”