Noroviruses: Laboratory Surrogates for Determining Survival and Inactivation
Dongyou Liu in Laboratory Models for Foodborne Infections, 2017
NoVs are small viruses about 27–32 nm in size and round in structure with an icosahedral symmetry. The human norovirus (HNoV) genome contains a single-stranded positive-sense RNA about 7.6 kb in length that is enclosed in a capsid without an envelope [3]. The capsid is made of 90 capsomers protruding from the shell that has 90 dimers of capsid protein. The genome has three open reading frames (ORFs). ORF1 (nucleotides 146–5359) is about 5 kb in size and encodes a ∼200 kDa nonstructural polyprotein. This nonstructural protein is cleaved to produce the N-terminal protein, the enzyme nucleoside triphosphatase, a 3A-like protein, a genome-linked viral protein (VpG), a 3C-like protease, and RNA-dependent RNA-polymerase (RdRp) [4]. ORF2 (nucleotides 5346–6935) is ∼1.8 kb in size and encodes the 57 kDa major structural capsid viral protein VP1; ORF3 (nucleotides 6938–7573) is ∼0.6 kb in size and encodes a small 22 kDa minor viral structural protein, VP2, reported to package the genome into virions [5,6]. The NoV genus at the time of this submission, is composed of five genogroups based on sequence analysis: genogroup I (GI) (prototype Norwalk virus); GII (prototype Snow Mountain virus); GIII (prototype bovine enteric calicivirus); GIV (prototype Alphatron and Ft. Lauderdale viruses); and GV (prototype Murine NoV) [7,8].
Exchange Factors
Juan Carlos Lacal, Frank McCormick in The ras Superfamily of GTPases, 2017
The GDS activity runs as two peaks on Mono Q FPLC columns. Both peaks (GDS-1 and -2) have a molecular weight of 53,000 on SDS-polyacrylamide gel electrophoresis and are indistinguishable by peptide map analysis. Partial amino acid sequence has now been obtained from purified smg p21 GDS and used to isolate cDNA clones.23 These reveal an open reading frame encoding a protein of 558 amino acids with a molecular weight 61,000. Expression of this protein in Escherichia coli confirms that it possesses guanine nucleotide exchange activity specific for smg p21/p21rap1. The primary structure of this GDS shows some low levels of homology to CDC25 and SDC25 protein and, in a different region, rather poorer homology to IRAI and NF-1 proteins, but not to GAP. The homology to CDC25/SDC25 is in the region of these proteins thought to possess the catalytic exchange activity toward yeast RAS proteins, but the homology to IRAI and NF-1 is outside the GTPase-activating catalytic domain. The significance of these homologies, particularly to IRAI and NF-1, has not yet been determined.
Determination
David Woolley, Adam Woolley in Practical Toxicology, 2017
Testing for genotoxicity acknowledges that there are basically two levels of effect–at the gene level and at the chromosome. At the former, mutations are sought that lead to localized changes at one or a few bases in the DNA, thereby changing the coding for the protein produced by the gene. A change from one base to another, or the misreading of a chemically altered base, may lead to a different amino acid being inserted into an otherwise normal protein; this is a point mutation. When a base or base pair is inserted or deleted, this is known as a frameshift mutation, as the reading frame of the code is changed, leading to an abnormal protein product. At the level of the chromosome, the changes are broadly in terms of structure or number; there may be changes in number due to effects on mitosis or meiosis and translocations, rearrangements, breaks, or gaps, which indicate an effect on the chromosomes themselves. DNA or chromosomal damage is detected directly or indirectly. Direct evidence comes from the induction of genetic change, such as the ability of Ames test bacteria to divide in the absence of a previously essential amino acid, or by examination of chromosomes in metaphase where breakages and abnormalities are evident under the microscope. Indirect evidence of genetic damage may be obtained by measurement of DNA repair in tissues. This is easier to detect in tissues that do not normally divide and is used in the assessment of unscheduled DNA synthesis (UDS) in hepatocytes.
Small, but mighty? Searching for human microproteins and their potential for understanding health and disease
Published in Expert Review of Proteomics, 2018
Annie Rathore, Thomas F. Martinez, Qian Chu, Alan Saghatelian
Microproteins are a rapidly expanding class of peptides and small proteins translated from protein-coding small open reading frames (smORFs, less than 100–150 codons in length). Microprotein is a term that refers to peptides and small proteins that are translated from smORFs and can include known genes. Microprotein discovery and characterization reshapes our understanding of proteome composition and reveals new biological pathways [1]. Genomes contain thousands of open reading frames (ORFs), defined as the protein-coding sequence between an in-frame start and stop codon. Annotation of protein-coding ORFs from DNA sequences became paramount as whole-genome sequencing projects reached completion [2]. Excellent computational methods were developed and utilized to define genes, but these tools needed to establish parameters to reduce false positives. For this reason, most genome annotation pipelines required ORFs to be at least 300 nucleotides long (i.e. 100 amino acids) resulting in most smORFs being missed [2]. To get an idea on the challenge of assigning protein-coding genes without a length cutoff, Basarai, Hieter, and Boeke identified ~260,000 smORFs between 2 and 99 codons when plotting all ORFs in the yeast genome [3]. Today, it is clear that smORFs and their corresponding microproteins make up a sizable fraction of the genome and proteome. As new genes, very little is known about the structure and function of microproteins making these genes an incredible opportunity for discovering new biology.
Challenges and promise at the interface of metaproteomics and genomics: an overview of recent progress in metaproteogenomic data analysis
Published in Expert Review of Proteomics, 2019
Henning Schiebenhoefer, Tim Van Den Bossche, Stephan Fuchs, Bernhard Y. Renard, Thilo Muth, Lennart Martens
The unbiased translation of sequencing data to protein sequences by using all possible reading frames is also a viable option. If information about the coding strand is available, sequencing data is translated in three reading frames (three-frame translation), otherwise in all six reading frames (six-frame translation). For the human genome, a six-frame translation would lead to a search database that is 70 times larger than the reference proteome from UniProtKB [24]. Related to that, proteogenomic databases often contain many obsolete sequences from reading frames that are not transcribed [63]. In the case of prokaryotes, the increase in database size will most likely be smaller, due to the greater gene density found in these organisms compared to humans. Still, the translated database would largely consist of nonsense protein sequences that are not present in the sample. The performance of searches on databases constructed this way has been shown to be comparable to, yet slightly worse than, databases based on gene prediction tools [33].
Recent advances in human norovirus research and implications for candidate vaccines
Published in Expert Review of Vaccines, 2020
Jordan E. Cates, Jan Vinjé, Umesh Parashar, Aron J. Hall
Noroviruses are classified within the Caliciviridae family, and have a 7.5 kb linear, positive sense, single-stranded RNA genome that is enclosed in a non-enveloped icosahedral capsid [24]. The genome is organized into three open reading frames (ORFs). ORF1 encodes a polyprotein that is co- and post-translationally cleaved into six non-structural viral proteins, including the RNA-dependent RNA polymerase (RdRp). ORF2 encodes VP1, the major structural capsid protein, which is composed of a shell (S) and two protruding (P) regions (P1 and P2). ORF3 encodes VP2, a minor structural capsid protein. Based on sequence differences of the VP1 protein, noroviruses are classified into at least ten genogroups (GI-GX), with most infections in humans caused by GI and GII viruses [25]. Noroviruses can be further classified into genotypes and P (polymerase)-types based on amino acid diversity of VP1 and nucleotide diversity of the RdRp region, respectively. Currently, there are at least 49 genotypes and 60 P-types [25]. Genogroup II genotype 4 (GII.4) viruses are further divided into epidemiologically important variants that carry the name of the location for the first strain from which the complete capsid sequence was submitted to GenBank (e.g., GII.4 Sydney).
Related Knowledge Centers
- Directionality
- DNA
- Genetic Code
- Molecular Biology
- Nucleic Acid
- Rna
- Amino Acid
- Nucleic Acid Sequence
- Translation
- Phosphoryl Group