Biology of microbes
Philip A. Geis in Cosmetic Microbiology, 2006
To understand the process in detail requires far more effort. In reality, at least seven enzymes are involved in the process described in the paragraph above: initiator protein, helicase, polymerases, repair nucleases, topoisomerase, single-strand DNA-binding proteins, and DNA ligase. The initiator protein first finds the right place to begin copying and guides the helicase to the correct position (an origin of replication site) on the nucleic acid. The helicase separates the DNA by breaking the weak bonds between the nucleotides to unwind the two strands of DNA. Then the polymerases arrive to join the free nucleotides to their matching complements on the old strands using the phosphate bond energy from the nucleotide to help form the new bond to the other nucleotides as they are added to the existing chain. These polymerases work along with primases that first synthesize a short (one to five nucleotides long) RNA primer. This primer allows DNA polymerase to begin catalyzing the addition of nucleotides to a new strand complementary to the existing template upon which the new DNA synthesis is based.
Regulation of Cell Functions
Enrique Pimentel in Handbook of Growth Factors, 2017
The enzyme DNA polymerase-α plays a key role in the replication of the eukaryotic genome. Regulation of human DNA polymerase-α gene expression occurs at the transcriptional level.233 During the mitogenic activation of quiescent cells, the steady-state RNA levels, rate of synthesis of nascent protein, and enzymatic activity of DNA polymerase-α exhibit a marked increase prior to the peak of DNA synthesis. Transcription of the DNA polymerase-α gene and other DNA replication-associated genes is reduced in quiescent cells but increases markedly upon mitogenic stimulation. However, the constitutive expression of DNA polymerase-α during the cell cycle suggests that a basal level of the enzyme is required for the maintenance of cellular DNA. In addition to DNA replication, DNA polymerase-α may have a role in DNA repair.
DNA-Binding Proteins and DNA-Synthesizing Enzymes in Eukaryotes
Lubomir S. Hnilica in Chromosomal Nonhistone Proteins, 2018
At least three different types of DNA polymerization can be classified: (1) replicative nuclear DNA synthesis, which is characterized by being semiconservative, symmetrical, bidirectional, and with short RNA-primed intermediates in the lagging strand; (2) repair DNA synthesis, occurring on both strands and restricted to short DNA gaps; (3) mitochondrial and adenovirus DNA synthesis, which is known to be continuous and proceeds in an asymmetrical way. DNA synthesis catalyzed by DNA polymerase α is a typical replicative type and is suggested to carry out replication of nuclear DNA. DNA polymerase α produces a short DNA fragment on ribo-primers in a quasiprocessive way. It may also be able to synthesize a longer DNA chain with the aid of other enzymes. On the contrary, DNA polymerase β works in a distributive way and incorporates nucleotides to the gaps on the activated DNA. Its properties suggest that DNA polymerase β performs a repair type of DNA synthesis. DNA polymerase α synthesizes DNA in vitro in a highly processive fashion. It can replicate mitochondrial and adenovirus DNAs, as well as synthesize the long, single-stranded DNA regions which might represent displaced strands of parental DNA being replicated asymmetrically. The details of the function of DNA polymerase α, β, and γ will be discussed later.
Interplay of Anti-Viral Vaccines with Biologic Agents and Immunomodulators in Individuals with Autoimmune and Autoinflammatory Diseases
Published in Immunological Investigations, 2021
Barbara E. Ostrov, Daniel Amsterdam
Inadequate response to vaccines can be attributed to rare occurrence of vaccine resistance. According to Kennedy and Read (Kennedy and Read 2018), documented cases of vaccine resistance are due to the inability of the vaccine to induce an immune response even when targeting multiple viral epitopes; inability to suppress pathogen multiplication within the host; and incapability of aborting transmission from vaccine protected hosts. Failure to protect against all serotypes of the target pathogen may cause insufficient response to a vaccine. In addition, mutations in viral target proteins can render vaccines less immunogenic. RNA viruses have exceedingly high mutation rates compared to DNA viruses because the enzyme required for polymerization – RNA polymerase – does not possess the proofreading capability of DNA polymerase. Given this fallibility, it is more challenging to produce consistently effective long-term vaccines for infections caused by RNA viruses such as influenza (with its well-known requirement of annual vaccine revisions) and notably, the recent concerns regarding the RNA virus which causes COVID-19, as well (Kennedy and Read 2020).
Plasma insulin-like growth factor binding protein 1 in pulmonary arterial hypertension
Published in Scandinavian Cardiovascular Journal, 2021
Habib Bouzina, Roger Hesselstrand, Göran Rådegran
Proseek multiplex cardiovascular II and III, as well as oncology II, 96-plex immunoassays (Olink Proteomics, Uppsala, Sweden) were used to analyse the present biomarkers. From these panels, biomarkers related to tumour biology were chosen for analysis; including cancer antigen 125 (CA-125), carbonic anhydrase 9 (CA-IX), carcinoembryonic antigen-related cell adhesion molecule (CEACAM)-1 and -5, cornulin, cyclin-dependent kinase inhibitor 1 (p21), folate receptor α (FR-α), human epididymis protein 4 (HE4), IGF-1 receptor, IGFBP-1, -2 and -7, kallikrein-6, -8, -11, -13 and -14, mesothelin, protein S100-A4 (S100A4) and -A11 (S100A11) and vimentin. These biomarkers, as well as N-terminal prohormone of brain natriuretic peptide (NT-proBNP), were assessed using proximity extension assays, as previously described [18,19]. In brief, oligonucleotide-labelled antibody pairs are used to detect targeted biomarkers, in order to avoid unspecific antibody binding and potential cross-reactivity events. As two related probes are brought in close proximity the oligonucleotides hybridise in a pair-wise manner. DNA polymerase addition results in a proximity-dependent DNA polymerisation event, creating a unique PCR target sequence. The DNA sequence is then detected and quantified using a microfluidic real-time PCR instrument (Biomark HD; Fluidigm, San Francisco, CA, USA). Data quality control and normalization is performed utilizing an internal extension control and an inter-plate control, in order to adjust for intra- and inter-run variation. Assay validation data and panel information are available at www.olink.com.
Potential therapeutic targets for Mpox: the evidence to date
Published in Expert Opinion on Therapeutic Targets, 2023
Siddappa N Byrareddy, Kalicharan Sharma, Shrikesh Sachdev, Athreya S. Reddy, Arpan Acharya, Kaylee M. Klaustermeier, Christian L Lorson, Kamal Singh
VACV DNA genome replication is conducted by a holoenzyme consisting of multiple proteins [25]. An essential component of this holoenzyme is E9, the DNA-dependent DNA polymerase belonging to B family DNA polymerases. The MPXV genome encodes F8L (OPG71) [2], also a B family DNA-dependent DNA polymerase [27], which shares ~ 98% identity with VACV E9. The first B family DNA polymerase (RB69) structure showed an overall architecture of this class of enzymes [28]. This structure showed a canonical polymerase domain consisting of the Thumb, Palm, and Fingers subdomains, as seen in the structure of the Klenow Fragment (KF) of E. coli DNA polymerase I [29]. A notable difference between these polymerases is the relative position of the 3’ − 5’ exonuclease domain, which is ~ 180° opposite to that in KF relative to the polymerase active site. Subsequent crystal structures of the RB69 polymerase showed that residues of a β-hairpin positioned in the major groove of the template-primer played a role in the partitioning of primer to the 3’ − 5’ exonuclease site upon mismatch nucleotide incorporation [28,30–32]. Indeed, a resistance mutation on topologically similar β-hairpin in poxviruses’ DNA polymerase showed the relevance of the resistance mechanism of nucleotide analogs mediated by 3’ − 5’ exonuclease function (discussed below).