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PubMed1997

Isolation of DNA from the centromere of human chromosome 7 by microdissection.

Centromeres remain the least characterized regions of human chromosomes because they have a very high content of repetitive DNA. Here, we describe a microdissection library from the centromeric region of human chromosome 7 and its use for generating sequence tagged sites (STSs). The library contains about 1500 clones with an average insert size of 150 bp and only about 15% of the clones harbour repetitive human DNA. Seven clones hybridizing to alphoid DNA were found to correspond to a fragment of the D7Z2 alphoid array on chromosome 7, thus confirming the origin of the library. A number of clones not containing known repetitive DNA were used to generate STSs that identified yeast artificial chromosomes (YACs) and in turn allowed the STSs to be placed on the physical map. One STS is located between the two Genethon genetic markers closest to the centromere on the q side. Another STS was located 3-4cM away in 7q11.2, while a third identified YACs containing both low-copy and alphoid sequences that are not yet mapped but are clearly centromeric. The library therefore comprises a collection of sequences from the centromeric region of chromosome 7 that can be used to generate STSs and to map the entire centromeric region.

Amniotic Fluid↗
PubMed2002

Non-carbonyl-stabilized metallocarbenoids in synthesis: the development of a tandem rhodium-catalyzed bamford-stevens/thermal aliphatic claisen rearrangement sequence.

A tandem rhodium-catalyzed Bamford-Stevens/Claisen rearrangement is presented. The tandem reaction uses Eschenmoser hydrazones for the in situ generation of non-carbonyl-stabilized diazo alkanes, which are presumably intercepted by Rh(II) catalysts to induce a 1,2-hydride migration. This sequence provides high levels of stereocontrol for the generation of simple acyclic (Z)-enol ethers. These enol ethers undergo either thermal or Lewis acid accelerated Claisen rearrangements to provide products of high diastereopurity. Also presented are cascade reactions, wherein a third chemical step occurs after the initial tandem sequence (i.e., Bamford-Stevens/Claisen/ene and Bamford-Stevens/Claisen/Cope).

Journal Article↗
PubMed1997

Identification of indispensable residues for specific DNA-binding in the imperfect tandem repeats of c-Myb R2R3.

The individual repeats, R2 and R3, of the minimum specific DNA-binding domain (R2R3) of c-Myb have very similar structures, with a helix-turn-helix variation motif, although their sequence identity in the tandem repeats is only 31%. From previous mutational and structural studies, the third helices in both repeats were shown to directly recognize the specific base sequence, PyAACG/TG. In order to elucidate the reason for the imperfection of the tandem repeats at amino acid positions other than the recognition helices, a series of R2R3 mutants was generated by swapping the helices and the N-terminus in R2 to those in R3. Consequently, the sequence composing the first helix of R2 was found to be essential for specific DNA-binding, in addition to the third recognition helix of R2. Further mutational studies revealed that the only indispensable residues in the first helix are Val103 and Val1O7, which are involved in the hydrophobic core of R2. These residues do not directly interact with the DNA, but they contribute to the correct formation of helix 1 and the characteristic packing of R2, which is slightly different from that of R3, and are required for specific base recognition through strong cooperativity with R3.

Binding Sites↗
PubMed1996

Isolation of a developmentally-regulated expressed sequence tag from bladder tissue using the mRNA differential display.

In order to gain insight into the molecular and cellular events that govern the structural and the functional properties in developing organs, we have conducted a study to identify genes that have a temporally-restricted expression in the bladder wall during fetal development. We utilized the mRNA differential display technique and compared the pattern of gene expression during the first, the second and the third trimester of gestation. We cloned and sequenced a cDNA fragment (bld-10) which was expressed during the second and third trimester but consistently absent during the first trimester. The bld-10 sequence is not related to any known gene in the GenBank database but has significant homology (89%) with human expressed sequence tag (EST) that has been cloned from human fetal heart and brain libraries. When used in Northern-blot hybridization as a probe, the fragment bld-10 generates two hybridization signals of 3.1 and 4.0 kb, that are minimally expressed during the first trimester of gestation and upregulated in the second and third trimester. Differential expression of this gene may be responsible for some of the profound changes which occur during organ development.

Animals↗
PubMed1999

Improved secondary structure predictions for a nicotinic receptor subunit: incorporation of solvent accessibility and experimental data into a two-dimensional representation.

Abstract A refined prediction of the nicotinic acetylcholine receptor (nAChR) subunits' secondary structure was computed with third-generation algorithms. The four selected programs, PHD, Predator, DSC, and NNSSP, based on different prediction approaches, were applied to each sequence of an alignment of nAChR and 5-HT3 receptor subunits, as well as a larger alignment with related subunit sequences from glycine and GABA receptors. A consensus prediction was computed for the nAChR subunits through a "winner takes all" method. By integrating the probabilities obtained with PHD, DSC, and NNSSP, this prediction was filtered in order to eliminate the singletons and to more precisely establish the structure limits (only 4% of the residues were modified). The final consensus secondary structure includes nine alpha-helices (24.2% of the residues, with an average length of 13.9 residues) and 17 beta-strands (22.5% of the residues, with an average length of 6.6 residues). The large extracellular domain is predicted to be mainly composed of beta-strands, with only two helices at the amino-terminal end. The transmembrane segments are predicted to be in a mixed alpha/beta topology (with a predominance of alpha-helices), with no known equivalent in the current protein database. The cytoplasmic domain is predicted to consist of two well-conserved amphipathic helices joined together by an unfolded stretch of variable length and sequence. In general, the segments predicted to occur in a periodic structure correspond to the more conserved regions, as defined by an analysis of sequence conservation per position performed on 152 superfamily members. The solvent accessibility of each residue was predicted from the multiple alignments with PHDacc. Each segment with more than three exposed residues was assumed to be external to the core protein. Overall, these data constitute an envelope of structural constraints. In a subsequent step, experimental data relative to the extracellular portion of the complete receptor were incorporated into the model. This led to a proposed two-dimensional representation of the secondary structure in which the peptide chain of the extracellular domain winds alternatively between the two interfaces of the subunit. Although this representation is not a tertiary structure and does not lead to predictions of specific beta-beta interaction, it should provide a basic framework for further mutagenesis investigations and for fold recognition (threading) searches.

Algorithms↗
PubMed1995

Binding of cephalothin and cefotaxime to D-ala-D-ala-peptidase reveals a functional basis of a natural mutation in a low-affinity penicillin-binding protein and in extended-spectrum beta-lactamases.

Two clinically-important beta-lactam antibiotics, cephalothin and cefotaxime, have been observed by X-ray crystallography bound to the reactive Ser62 of the D-alanyl-D-alanine carboxypeptidase/transpeptidase of Streptomyces sp. R61. Refinement of the two crystal structures produced R factors for 3 sigma (F) data of 0.166 (to 1.8 A) and 0.170 (to 2.0 A) for the cephalothin and cefotaxime complexes, respectively. In each complex, a water molecule is within 3.1 and 3.6 A of the acylated beta-lactam carbonyl carbon atom, but is poorly activated by active site residues for nucleophilic attack and deacylation. This apparent lack of good stereochemistry for facile hydrolysis is in accord with the long half-lives of cephalosporin intermediates in solution (20-40 h) and the efficacy of these beta-lactams as inhibitors of bacterial cell wall synthesis. Different hydrogen binding patterns of the two cephalosporins to Thr301 are consistent with the low cefotaxime affinity of an altered penicillin-binding protein, PBP-2x, reported in cefotaxime-resistant strains of Streptococcus pneumoniae, and with the ability of mutant class A beta-lactamases to hydrolyze third-generation cephalosporins.

Amino Acid Sequence↗
PubMed1994

An immunodominant epitope in a functional domain near the N-terminus of human granulocyte-macrophage colony-stimulating factor identified by cross-reaction of synthetic peptides with neutralizing anti-protein and anti-peptide antibodies.

We produced polyclonal and monoclonal antibodies (MAbs) against recombinant human (rh) granulocyte-macrophage colony-stimulating factor (GM-CSF) and performed studies of epitope mapping by ELISA, using five synthetic peptides corresponding to sequences along this molecule. Additionally, anti-peptide MAbs were generated. The antibody ability to inhibit rhGM-CSF activity was determined using as bioassay the MO7e cell line, which is dependent on hGM-CSF for growth in vitro. An immunodominant epitope able to induce the highest neutralization antibody titers was identified near the N terminus of hGM-CSF. A synthetic peptide 14-24, homologous to a sequence including part of the first alpha-helix of the molecule, was recognized by neutralizing anti-protein antibodies. Similarly, MAbs anti- 14-24 cross-reacted with rhGM-CSF and specifically blocked its function. Replacement of Val16 or Asn17 with alanine greatly reduced the antibody-binding capacity to peptide 14-24, whereas substitution of Gln20 or Glu21 was less critical. Monoclonal antibodies generated against residues 30-41 (corresponding to an intrahelical loop) and 79-91 (homologous to a sequence including part of the third alpha-helix) or its analog [Ala88](79-91)beta Ala-Cys, were conformation dependent and nonneutralizing: they failed to react or bound poorly to rhGM-CSF in ELISA, but readily recognized the homologous sequence in the denatured protein, by Western blotting.

Amino Acid Sequence↗
PubMed2000

A phylogenetic tree of the Wnt genes based on all available full-length sequences, including five from the cephalochordate amphioxus.

The WNT: gene family is large, and new members are still being discovered. We constructed a parsimony tree for the WNT: family based on all 82 of the full-length sequences currently available. The inclusion of sequences from the cephalochordate amphioxus is especially useful in comprehensive gene trees, because the amphioxus genes in each subfamily often mark the base of the vertebrate diversification. We thus isolated full-length cDNAs of five amphioxus WNT: genes (AmphiWnt1, AmphiWnt4, AmphiWnt7, AmphiWnt8, and AmphiWnt11) for addition to the overall WNT: family tree. The analysis combined amino acid and nucleotide sequences (excluding third codon positions), taking into account 97% of the available data for each sequence. This combinatorial method had the advantage of generating a single most-parsimonious tree that was trichotomy-free. The reliability of the nodes was assessed by both jackknifing and Bremer support (decay index). A regression analysis revealed that branch length was strongly correlated with branch support, and possible reasons for this pattern are discussed. The tree topology suggested that in amphioxus, at least an AmphiWnt5 and an AmphiWnt10 have yet to be discovered.

Animals↗
PubMed1999

Dopamine sulphate: an enigma resolved.

1. The source and physiological significance of dopamine (DA) sulphate, which exists in plasma at much higher concentrations than free DA, have long been a puzzle. The present article reviews how the convergence of modern molecular and traditional clinical approaches is shedding new light on the origins and meaning of DA sulphate. 2. The sulphotransferase isoenzyme responsible for production of DA sulphate in humans (SULT1A3) has been cloned and shown to be expressed in large quantities in the gastro-intestinal tract, but not in liver. No orthologue of SULT1A3 has yet been identified in other species, consistent with the greater importance of sulphate conjugation of DA in humans than in most animals. 3. Diet has a major impact on plasma DA sulphate, with dramatic increases after ingestion of meals and foods rich in biogenic amines; however, substantial amounts of DA sulphate remaining after prolonged fasting indicate the presence of a mainly endogenous source. The lack of influence of acute or chronic changes in sympathetic outflow or of sympathoneural degeneration on plasma DA sulphate indicates that DA sulphate does not derive from sympathetic nerve. Relatively low rates of production from intravenously infused DA indicate that very little DA sulphate (< 2%) derives from metabolism of circulating DA, such as in red cells or platelets. 4. Consistent increments in DA sulphate from arterial to the outflowing venous plasma draining mesenteric organs, without increments across other organs or tissues (e.g., heart, lungs, liver), indicate that the gastrointestinal tract is a major source of more than 75% of DA sulphate produced in the body. The gastro-intestinal tract is also the site of a novel DA autocrine/paracrine system that produces nearly 50% of the DA in the body. Therefore, production of DA sulphate appears to reflect an enzymatic 'gut-blood' barrier for detoxifying dietary biogenic amines and delimiting autocrine/paracrine effects of endogenous DA generated in a novel 'third catecholamine system'.

Base Sequence↗
PubMed1989

Evolutionary pathways of the calcitonin (CALC) genes.

Recombinant DNA techniques have made it possible to establish the structure of various genes encoding polypeptide hormones. Comparison of nucleotide sequences of the calcitonin (CALC) genes in man has revealed surprising similarities and variations. These findings and the homologies among the sequences in different species offered an opportunity for speculation about relationships between these genes and about their evolutionary origin. The first gene (CALC-I) directing the synthesis of calcitonin (CT) or CT gene-related peptide (CGRP) comprises six exons and gives rise to two mRNAs by an alternative RNA-processing mechanism. The homology between CGRP and CT reflects their common origin. The human genome contains a second gene (CALC-II) that is structurally related to the CALC-I gene. The CALC-II RNA transcripts do not appear to be differentially processed, as only preproCGRP-II mRNA and not preproCT-II is detected. The first and second CT/CGRP genes probably have evolved from a common ancestor gene early in evolution. Meanwhile, a third genomic locus containing nucleotide sequences highly homologous to exons 2 and 3 of both CALC genes was detected and probably generated by duplication of a part of CALC-II. This locus is not likely to encode a CT- or CGRP-related polypeptide hormone. The CALC genes and this last (pseudo) gene are located on the short arm of chromosome 11. Recently, islet- or insulinoma-amyloid polypeptide (IAPP) was isolated as a major constituent of amyloid present in human insulinoma and in pancreatic islet amyloid in noninsulin-dependent diabetes mellitus. IAPP shows 46% amino acid sequence homology with human CGRP-II.(ABSTRACT TRUNCATED AT 250 WORDS)

Amyloid↗
PubMed1995

Novel system for analysis of group I 3' splice site reactions based on functional trans-interaction of the P1/P10 reaction helix with the ribozyme's catalytic core.

A group I intron from a bacterial tRNA precursor has been converted into an RNA enzyme that catalyzes the efficient polymerization of oligoribonucleotide analogs of tRNA exons using a reaction scheme consisting of multiple cycles of reverse and forward exon ligation reactions. Here, we present results showing that this system represents a novel and useful tool for the analysis of 3' splice site reactions of group I ribozymes. First, analysis of variant substrates containing base substitutions in group I secondary structure elements P1, P9.0 and P10 confirms that exon polymerization is dependent on these structures, and therefore constitutes an appropriate and relevant model system for studying the exon ligation step of splicing. Second, to probe interactions between the intron's catalytic core and the bases and backbone of the P1/P10 reaction helix, two successful strategies for separating the internal guide sequence from the intron core were devised. One such strategy uses a construct in which the reaction helix interacts functionally with the catalytic core using only tertiary contacts. Further stabilization of this interaction through the inclusion of a 7 bp intermolecular P2 helix generates increased reaction efficiency. Third, when provided with two reaction helices, the ribozyme synthesizes mixed polymers through a mechanism that involves sequential binding and release of the duplexes. Fourth, in these reactions, turnover of the external guide sequence requires unwinding and annealing of the P2 helix, suggesting that P2 unwinding may occur during group I splicing. These results provide novel experimental tools to probe the relatively poorly understood 3' splice site reactions of group I introns, and may be relevant to ribozyme-catalyzed assembly and recombination of oligomers in prebiotic scenarios.

Base Sequence↗
PubMed1988

Proteolytic processing of pro-alpha and pro-beta precursors from human beta-hexosaminidase. Generation of the mature alpha and beta a beta b subunits.

There are two major isozymes of human lysosomal beta-hexosaminidase (beta-N-acetylhexosaminidase, EC 3.2.1.52), hexosaminidase A, alpha(beta a beta b), and hexosaminidase B, 2(beta a beta b). The alpha subunit contains a single polypeptide chain, while the beta subunit is composed of two nonidentical chains (beta a and beta b) derived from a common pro-beta precursor. The mature subunits, like those of most lysosomal enzymes, are produced through the proteolytic processing of propolypeptides once they enter the lysosome. In order to define the structure of the alpha and beta subunits generated in the lysosome, the alpha, beta a, and beta b polypeptides of hexosaminidase A and B were separated by a combination of molecular sieve and ion exchange high performance liquid chromatography, and amino-terminal sequences were determined. These were localized to the deduced amino acid sequences of previously isolated cDNAs coding for the prepro-alpha and beta polypeptides. From this analysis, the sites of hydrolysis generating the mature alpha, beta a, and beta b chains from hexosaminidase A and B could be determined. First, the signal peptide, required for processing of the pre-propolypeptides through the rough endoplasmic reticulum was predicted from the first in-frame Met residue on the cDNA. Second, amino acid sequencing defined the amino termini of the mature polypeptide chains and identified the pro-sequences removed from both the pro-alpha and pro-beta polypeptides. Third, an internal cleavage resulted in the removal of a tetrapeptide, Arg-Gln-Asn-Lys, and tripeptide, Arg-Gln-Asn, from the pro-beta chain of hexosaminidase A and B, respectively , to generate the beta b and beta a chains. This result localized the beta b and beta a chains to the amino-terminal and carboxyl-terminal halves of the pro-beta sequence, respectively. Finally, we previously reported minimal or no carboxyl-terminal processing of the pro-beta chain in the lysosome. On the other hand, we suggest that there is trimming at the carboxyl terminus of the pro-alpha chain based on comparison of molecular weights of deglycosylated alpha with the isolated beta b and beta a chains comprising the mature beta subunit with those predicted from the cDNA. Thus, in the lysosome the pro forms of hexosaminidase A and B undergo extensive proteolytic processing which, while specific in nature, has the appearance of removing easily accessible, nonessential domains, rather than contributing to biosynthetic maturation of function.

Amino Acid Sequence↗
PubMed1994

Human erythrocyte ankyrin, a cytoskeleton component, generates the p57 membrane proteinase which cleaves C3, the third component of complement.

Human erythrocytes express a p57 membrane serine proteinase which cleaves C3, the third component of complement. Amino acid analysis of the first 20 N-terminal residues of the purified p57 proteinase demonstrated 100% homology with residues 910-929 of erythrocyte ankyrin, a sequence localized in its Mr = 62 kDa fragment. Thus, we analyzed whether ankyrin could generate the p57 C3-cleaving activity. We demonstrate herein that: 1) anti-ankyrin antibodies react with purified p57 proteinase; 2) anti-p57 proteinase antibodies react with purified ankyrin but not with spectrin, another cytoskeleton component; 3) while purified ankyrin did not carry any detectable proteinase activity, limited proteolysis of ankyrin by immobilized chymotrypsin generated this C3-cleaving serine proteinase. Spectrin did not generate any C3-cleaving activity. This is the first demonstration that erythrocyte ankyrin could generate a proteinase activity, which in addition, cleaves specifically human C3.

Amino Acid Sequence↗
PubMed1998

The Human Genome Project: from mapping to sequencing.

Until recently, the "human genome" programs were mainly directed towards the development of maps to identify disease genes. The genetic map comprises about 8000 highly informative second generation markers of the microsatellite type. The density of markers is now sufficient to localize a gene for a monogenic disease with a precision of 1 to 2 million base pairs easily, and to define intervals which contain susceptibility genes for multifactorial disorders. A third generation map based on single nucleotide polymorphisms that can be genotyped using DNA chip technology is in progress. The physical map, based on sets of overlapping yeast artificial chromosomes ordered using sequence-tagged sites, covers over 90% of the genome. However, this physical map cannot serve as a support for sequencing because of the numerous rearrangements that occur in yeast artificial chromosomes. An international network of laboratories has mapped a set of more than 30,000 expressed sequences from cDNAs using whole genome radiation hybrids that enable integration of genes within existing maps. The human genome program is now progressively shifting to massive sequencing, although sequence ready maps are not available for the major part of the human genome. Similarly, our capacity to interpret the available genomic sequence remains limited.

Genetic Linkage↗
PubMed1989

Molecular characterization of a major autoantibody-associated cross-reactive idiotype in Sjogren's syndrome.

Primary Sjogren's syndrome is an autoimmune disorder characterized by lymphocytic infiltration of the salivary and lacrimal glands, producing associated dry eyes (keratoconjunctivitis sicca), dry mouth, and intermittently swollen salivary glands. A high proportion of the infiltrating B lymphocytes express surface and cytoplasmic Ig bearing a kappa-L chain-associated CRI defined by reactivity with the murine mAb, 17.109. To determine the structural basis for CRI expression in this disease, we generated CRI+ lymphoblastoid cell lines and a cDNA library from lymphocytes extracted from Sjogren's syndrome patients' salivary gland biopsy specimens. Nucleic acid sequence analyses of the mRNA of one such 17.109-CRI+ lymphoblastoid cell line (NOV) reveals the expressed kappa light chain variable region gene (V kappa gene) to be homologous to Humkv325, a conserved V kappa gene used at relatively high frequency in certain B cell malignancies. In addition, synthetic oligonucleotides, corresponding to the first and third frameworks and the second complementarity determining region of the Humkv325 gene, were used to identify and isolate clones from a cDNA library generated from SS salivary gland lymphocytes. Clones annealing specifically with one or more of these oligonucleotide probes contained kappa light chain cDNA. The sequences corresponding to the variable region of two clones (Taykv320 and Taykv306) were homologous to Humkv325. The V kappa genes of four other cDNA clones (Taykv322, Taykv310, Taykv308, and Taykv312) most likely were generated somatically from the rearranged Humkv325 gene through a limited number of nucleic acid base substitutions. Our results suggest that the high frequency of 17.109-CRI expression in Sjogren's syndrome patients results from a multiclonal expansion of B cells using Humkv325, and that the expressed Humkv325 may undergo somatic diversification in an apparent Ag-driven response.

Amino Acid Sequence↗
PubMed1997

Antigenic variation of core, NS3, and NS5 proteins among genotypes of hepatitis C virus.

Assays that detect antibody to hepatitis C virus (HCV) are used to screen blood donors and patients with hepatitis. Current enzyme-linked immunosorbent assay (ELISA)-based methods are invariably based upon antigens from expressed recombinant proteins or oligopeptides from HCV type 1. Some HCV antigens used in screening assays are coded by regions of the HCV genome that show extensive variability; therefore, HCV type 1-based assays may be less effective for the detection of antibody elicited by infection with other genotypes. In this study, we have measured antibody reactivity of sera from 110 hepatitis C patients infected with type 1b, 3a, or 4a to genotype-specific and cross-reactive epitopes present in recombinant proteins from HCV genotypes 1b (core, NS3, and NS5), 3a (NS3, NS5), and 4a (core, NS3), corresponding to those used in current third-generation screening ELISAs. By comparing the serological reactivities of sera to type-homologous and type-heterologous antigens, we detected a significant type-specific component to the reactivity to NS3 (61 to 77% of the total reactivity) and NS5 (60% of the total reactivity). Furthermore, despite the similarities in the amino acid sequences of the core antigens of type 1b and type 4a, we also found significantly greater reactivity to type-homologous antigens, with approximately 25% of reactivity being type specific. These findings are consistent with previous findings of fivefold weaker reactivity of sera from HCV type 2- and HCV type 3-infected blood donors in the currently used third-generation ELISAs and suggest that these assays are suboptimal for screening populations in which the predominant genotype is not type 1.

Amino Acid Sequence↗
PubMed1998

Sequence analysis of the mip gene of the soilborne pathogen Legionella longbeachae.

To understand the basis of pathogenesis by Legionella longbeachae serogroup 1, the importance of the Mip protein in this species was examined. Amino-terminal analysis of the purified, cloned L. longbeachae serogroup 1 ATCC 33462 Mip protein confirmed that the cloned gene protein was expressed and processed in an Escherichia coli background. DNA sequence analysis of plasmid pIMVS27, containing the entire L. longbeachae serogroup 1 mip gene, revealed a high degree of homology to the mip gene of Legionella pneumophila serogroup 1, 76% homology at the DNA level and 87% identity at the amino acid level. Primer extension analysis determined that the start site of transcription was the same for both species, with some differences observed for the -10 and -35 promoter regions. Primers designed from the mip gene sequence obtained for L. longbeachae serogroup 1 ATCC 33462 were used to amplify the mip genes from L. longbeachae serogroup 2 ATCC 33484 and an Australian clinical isolate of L. longbeachae serogroup 1 A5H5. The mip gene from A5H5 was 100% identical to the type strain sequence. The serogroup 2 strain of L. longbeachae differed by 2 base pairs in third-codon positions. Allelic exchange mutagenesis was used to generate an isogenic mip mutant in ATCC 33462 and strain A5H5. The ATCC mip mutant was unable to infect a strain of Acanthamoebae sp. both in liquid and in a potting mix coculture system, while the A5H5 mip mutant behaved in a manner siilar to that of L. pneumophila serogroup 1, i.e., it displayed a reduced capacity to infect and multiply within Acanthamoebae. To determine if this mutation resulted in reduced virulence in the guinea pig animal model, the A5H5 mip mutant and its parent strain were assessed for their abilities to establish an infection after aerosol exposure. Unlike the virulent parent strain, the mutant strain did not kill any animals under two different dose regimes. The data indicate that the Mip protein plays an important role in the intracellular life cycle of L. longbeachae serogroup 1 species and is required for full virulence.

Amino Acid Sequence↗