Crystal structure of aThermus aquaticusdiversity-generating retroelement variable protein

AbstractDiversity-generating retroelements (DGRs) are widely distributed in bacteria, archaea, and microbial viruses, and bring about unparalleled levels of sequence variation in target proteins. While DGR variable proteins share low sequence identity, the structures of several such proteins have revealed the C-type lectin (CLec)-fold as a conserved scaffold for accommodating massive sequence variation. This conservation has led to the suggestion that the CLec-fold may be useful in molecular surface display applications. Thermostability is an attractive feature in such applications, and thus we studied the variable protein of a DGR encoded by the thermophileThermus aquaticus. We report here the 2.8 Å resolution crystal structure of the variable protein from theT. aquaticusDGR, called TaqVP, and confirm that it has a CLec-fold. Remarkably, its variable region is nearly identical in structure to those of several other CLec-fold DGR variable proteins despite low sequence identity among these. TaqVP was found to be thermostable, which appears to be a property shared by several CLec-fold DGR variable proteins. These results provide impetus for the pursuit of the DGR variable protein CLec-fold in molecular display applications.

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Structure of the N-terminal dimerization domain of CEACAM7

Acta Crystallographica Section F Structural Biology Communications ◽

10.1107/s2053230x15013576 ◽

2015 ◽

Vol 71 (9) ◽

pp. 1169-1175 ◽

Cited By ~ 1

Author(s):

Daniel A. Bonsor ◽

Dorothy Beckett ◽

Eric J. Sundberg

Keyword(s):

Crystal Structure ◽

Hydrogen Bond ◽

Sequence Variation ◽

Cellular Adhesion ◽

Sedimentation Equilibrium ◽

Colorectal Cancers ◽

Dimerization Domain ◽

Adhesion Protein ◽

Dimerization Interface ◽

Colon And Rectum

CEACAM7 is a human cellular adhesion protein that is expressed on the surface of colon and rectum epithelial cells and is downregulated in colorectal cancers. It achieves cell adhesion through dimerization of the N-terminal IgV domain. The crystal structure of the N-terminal dimerization domain of CEACAM has been determined at 1.47 Å resolution. The overall fold of CEACAM7 is similar to those of CEACAM1 and CEACAM5; however, there are differences, the most notable of which is an insertion that causes theC′′ strand to buckle, leading to the creation of a hydrogen bond in the dimerization interface. TheKdimerizationfor CEACAM7 determined by sedimentation equilibrium is tenfold tighter than that measured for CEACAM5. These findings suggest that the dimerization affinities of CEACAMs are modulatedviasequence variation in the dimerization surface.

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Structural basis for UFM1 transfer from UBA5 to UFC1

Nature Communications ◽

10.1038/s41467-021-25994-6 ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Manoj Kumar ◽

Prasanth Padala ◽

Jamal Fahoum ◽

Fouad Hassouna ◽

Tomer Tsaban ◽

...

Keyword(s):

Crystal Structure ◽

Active Site ◽

Structural Basis ◽

Adenylation Domain ◽

Post Translational Modification ◽

Target Proteins ◽

Cellular Processes ◽

Enzyme Cascade ◽

Linear Sequence ◽

C Terminus

AbstractUfmylation is a post-translational modification essential for regulating key cellular processes. A three-enzyme cascade involving E1, E2 and E3 is required for UFM1 attachment to target proteins. How UBA5 (E1) and UFC1 (E2) cooperatively activate and transfer UFM1 is still unclear. Here, we present the crystal structure of UFC1 bound to the C-terminus of UBA5, revealing how UBA5 interacts with UFC1 via a short linear sequence, not observed in other E1-E2 complexes. We find that UBA5 has a region outside the adenylation domain that is dispensable for UFC1 binding but critical for UFM1 transfer. This region moves next to UFC1’s active site Cys and compensates for a missing loop in UFC1, which exists in other E2s and is needed for the transfer. Overall, our findings advance the understanding of UFM1’s conjugation machinery and may serve as a basis for the development of ufmylation inhibitors.

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Kinetics of Genetic Variation of the Mycoplasma genitalium MG192 Gene in Experimentally Infected Chimpanzees

Infection and Immunity ◽

10.1128/iai.01162-15 ◽

2015 ◽

Vol 84 (3) ◽

pp. 747-753 ◽

Cited By ~ 10

Author(s):

Liang Ma ◽

Jørgen S. Jensen ◽

Miriam Mancuso ◽

Leann Myers ◽

David H. Martin

Keyword(s):

Sequence Analysis ◽

Sequence Variation ◽

Variable Region ◽

Mycoplasma Genitalium ◽

Immune Selection ◽

Content Type ◽

Time Points ◽

Single Colony ◽

Nucleotide Sequence Variation ◽

Kinetics Of

Mycoplasma genitalium, a human pathogen associated with sexually transmitted diseases, is capable of causing chronic infections, though mechanisms for persistence remain unclear. Previous studies have found that variation of the MgPa operon occurs by recombination of repetitive chromosomal sequences (known as MgPars) into the MG191 and MG192 genes carried on this operon, which may lead to antigenic variation and immune evasion. In this study, we determined the kinetics of MG192 sequence variation during the course of experimental infection using archived specimens from two chimpanzees infected withM. genitaliumstrain G37. The highly variable region of MG192 was amplified by PCR fromM. genitaliumisolates obtained at various time points postinfection (p.i.). Sequence analysis revealed that MG192 sequence variation began at 5 weeks p.i. With the progression of infection, sequence changes accumulated throughout the MG192 variable region. The presence of MG192 variants at specific time points was confirmed by variant-specific PCR assays and sequence analysis of single-colony clonedM. genitaliumorganisms. MG192 nucleotide sequence variation correlated with estimated recombination events, predicted amino acid changes, and time of seroconversion, a finding consistent with immune selection of MG192 variants. In addition, we provided evidence that MG192 sequence variation occurred during the process ofM. genitaliumsingle-colony cloning. Such spontaneous variation suggests that some MG192 variation is independent of immune selection but may form the basis for subsequent immune selection.

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The X-Ray Crystal Structure of the Keratin 1-Keratin 10 Helix 2B Heterodimer Reveals Molecular Surface Properties and Biochemical Insights into Human Skin Disease

Journal of Investigative Dermatology ◽

10.1016/j.jid.2016.08.018 ◽

2017 ◽

Vol 137 (1) ◽

pp. 142-150 ◽

Cited By ~ 23

Author(s):

Christopher G. Bunick ◽

Leonard M. Milstone

Keyword(s):

Crystal Structure ◽

Surface Properties ◽

Human Skin ◽

Skin Disease ◽

Molecular Surface ◽

X Ray ◽

Keratin 1

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Divergent architecture of the heterotrimeric NatC complex explains N-terminal acetylation of cognate substrates

Nature Communications ◽

10.1038/s41467-020-19321-8 ◽

2020 ◽

Vol 11 (1) ◽

Cited By ~ 1

Author(s):

Stephan Grunwald ◽

Linus V. M. Hopf ◽

Tobias Bock-Bierbaum ◽

Ciara C. M. Lally ◽

Christian M. T. Spahn ◽

...

Keyword(s):

Crystal Structure ◽

Saccharomyces Cerevisiae ◽

Amino Acids ◽

Essential Role ◽

Catalytic Mechanism ◽

Detailed Structure ◽

Auxiliary Subunits ◽

Target Proteins ◽

Ribosome Binding ◽

Specific Ligand

Abstract The heterotrimeric NatC complex, comprising the catalytic Naa30 and the two auxiliary subunits Naa35 and Naa38, co-translationally acetylates the N-termini of numerous eukaryotic target proteins. Despite its unique subunit composition, its essential role for many aspects of cellular function and its suggested involvement in disease, structure and mechanism of NatC have remained unknown. Here, we present the crystal structure of the Saccharomyces cerevisiae NatC complex, which exhibits a strikingly different architecture compared to previously described N-terminal acetyltransferase (NAT) complexes. Cofactor and ligand-bound structures reveal how the first four amino acids of cognate substrates are recognized at the Naa30–Naa35 interface. A sequence-specific, ligand-induced conformational change in Naa30 enables efficient acetylation. Based on detailed structure–function studies, we suggest a catalytic mechanism and identify a ribosome-binding patch in an elongated tip region of NatC. Our study reveals how NAT machineries have divergently evolved to N-terminally acetylate specific subsets of target proteins.

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Crystal Structure of Human Profilaggrin S100 Domain and Identification of Target Proteins Annexin II, Stratifin, and HSP27

Journal of Investigative Dermatology ◽

10.1038/jid.2015.102 ◽

2015 ◽

Vol 135 (7) ◽

pp. 1801-1809 ◽

Cited By ~ 8

Author(s):

Christopher G. Bunick ◽

Richard B. Presland ◽

Owen T. Lawrence ◽

David J. Pearton ◽

Leonard M. Milstone ◽

...

Keyword(s):

Crystal Structure ◽

Annexin Ii ◽

Target Proteins

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Purification, crystallization and preliminary crystallographic analysis of the full-length cystathionine β-synthase fromApis mellifera

Acta Crystallographica Section F Structural Biology and Crystallization Communications ◽

10.1107/s1744309112038638 ◽

2012 ◽

Vol 68 (11) ◽

pp. 1323-1328 ◽

Cited By ~ 3

Author(s):

Iker Oyenarte ◽

Tomas Majtan ◽

June Ereño ◽

María Angeles Corral-Rodríguez ◽

Jaroslav Klaudiny ◽

...

Keyword(s):

Crystal Structure ◽

Crystallographic Analysis ◽

Full Length ◽

Asymmetric Unit ◽

Connective Tissues ◽

Unit Cell Parameters ◽

Cell Parameters ◽

Sequence Identity ◽

Transsulfuration Pathway ◽

Multisystemic Disease

Cystathionine β-synthase (CBS) is a pyridoxal-5′-phosphate-dependent enzyme that catalyzes the first step of the transsulfuration pathway, namely the condensation of serine with homocysteine to form cystathionine. Mutations in the CBS gene are the single most common cause of hereditary homocystinuria, a multisystemic disease affecting to various extents the vasculature, connective tissues and central nervous system. At present, the crystal structure of CBS fromDrosophila melanogasteris the only available structure of the full-length enzyme. Here we describe a cloning, overexpression, purification and preliminary crystallographic analysis of a full-length CBS fromApis mellifera(AmCBS) which maintains 51 and 46% sequence identity with itsDrosophilaand human homologs, respectively. TheAmCBS yielded crystals belonging to space groupP212121, with unit-cell parametersa= 85.90,b= 95.87,c= 180.33 Å. Diffraction data were collected to a resolution of 3.0 Å. The crystal structure contained two molecules in the asymmetric unit which presumably correspond to the dimeric species observed in solution.

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Allotypically related sequences in the Fd fragment of rabbit immunoglobulin heavy chains

Biochemical Journal ◽

10.1042/bj1240301 ◽

1971 ◽

Vol 124 (2) ◽

pp. 301-318 ◽

Cited By ~ 85

Author(s):

L. E. Mole ◽

S. A. Jackson ◽

R. R. Porter ◽

J. M. Wilkinson

Keyword(s):

Heavy Chain ◽

Immunoglobulin G ◽

Sequence Variation ◽

Variable Region ◽

Related Sequence ◽

Heavy Chains ◽

Rabbit Immunoglobulin ◽

Variable Section ◽

Terminal Section ◽

Constant Section

The sequence has been completed of the N-terminal 94 residues of the variable section of the Fd fragment of heavy chains from rabbit immunoglobulin G (IgG) of allotype As1. Most of the sequence of the same section from IgG of allotype Aa3 is also reported. These results, in conjunction with a substantial sequence of the variable region of allotype Aa2 reported elsewhere (Fleischman, 1971), show the presence of 16 positions (including six consecutive positions) in which the residue present correlates with the allotype. No allotype-related sequence variation has been found in the constant section of the Fd fragment. This evidence supports the view that two genes code for the heavy chain and it can be used as evidence in favour of somatic mutation as the origin of the variability in the sequence of the N-terminal section. The evolutionary origin of the ‘a’ locus allotypes of rabbit immunoglobulins remains obscure.

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