Studies of ribosomal proteins have given insight to the function and structure of the ribosome. During ribosome assembly, individual ribosomal proteins that initially recognize rRNA must bind to the assembly particle in a certain order and work cooperatively, rather than independently (1). Two of these important ribosomal proteins are S7 and S8. Detailed study of these and other ribosomal proteins is vital to the understanding of protein synthesis.Here we report the structure, chromosome location and complete sequence of a cDNA in the mosquito Anopheles gambiae encoding a homologue of the human ribosomal S7 protein. The clone was isolated from a cDNA library constructed using poly(A+) RNA of adult female and male mosquitoes from the G3 strain of A.gambiae (2). The complete 817 nucleotidelong cDNA contains a continuous open reading frame of 193 codons which potentially encodes a peptide with a molecular weight of 22206.90 Daltons and an isoelectric point of 9.945. Furthermore, this peptide shows a high level of basic amino acids (37), consistent with characterizations of S7 and S8-like proteins in other organisms (3). A presumed polyadenylation signal, AATAAA is located at bases 771-776.At the amino acid level, the open reading frame shows 71.7% identity with human ribosomal protein S7; 70.6% identity with Xenopus laevis ribosomal protein S8, and 61.7% identity with rat ribosomal protein S8 (Figure 1). Nucleotide comparison of the A.gambiae ribosomal protein cDNA coding sequence reveals a 66.7% identity with human ribosomal protein S7, but there are no evident similarities among the 5' or 3' non-coding sequences.In situ hybridization to the ovarian nurse cell polyrene chromosomes ofA.gambiae revealed its location at division 38B in the A.gambiae genome.
Anopheles gambiae, the primary vector of human malaria in Africa, is responsible for approximately a million deaths per year, mostly of children. Despite its significance in disease transmission, this mosquito has not been studied extensively by genetic or molecular techniques. To facilitate studies on this vector, a genetic map has been developed that covers the X chromosome at an average resolution of 2 centimorgans. This map has been integrated with the chromosome banding pattern and used to localize a recessive, sex-linked mutation (white eye) to within 1 centimorgan of flanking markers.
High molecular weight serpins are members of a large superfamily of structurally conserved proteins that inactivate target proteinases by a suicide substrate-like mechanism. In vertebrates, different clades of serpins distribute predominantly to either the intracellular or extracellular space. Although much is known about the function, structure, and inhibitory mechanism of circulating serpins such as ␣ 1 -antitrypsin (SERPINA1) and antithrombin III (SERPINC1), relatively little is known about the function of the vertebrate intracellular (clade B) serpins. To gain a better understanding of the biology of the intracellular serpins, we initiated a comparative genomics study using Caenorhabditis elegans as a model system. A screen of the C. elegans genomic and cDNA databases revealed nine serpin genes, tandemly arrayed on chromosome V. Although the C. elegans serpins represent a unique clade (L), they share significant functional homology with members of the clade B group of intracellular serpins, since they lack typical N-terminal signal peptides and reside intracellularly. To determine whether nematode serpins function as proteinase inhibitors, one family member, srp-2, was chosen for further characterization. Biochemical analysis of recombinant SRP-2 protein revealed SRP-2 to be a dual cross-class inhibitor of the apoptosis-related serine proteinase, granzyme B, and the lysosomal cysteine proteinases, cathepsins K, L, S, and V. Analysis of temporal and spatial expression indicated that SRP-2 was present during early embryonic development and highly expressed in the intestine and hypoderm of larval and adult worms. Transgenic animals engineered to overexpress SRP-2 were slow growing and/or arrested at the first, second, or third larval stages. These data suggest that perturbations of serpin-proteinase balance are critical for correct postembryonic development in C. elegans.Serpins are a unique class of proteinase inhibitors that irreversibly neutralize target proteinases by a mechanism involving the conformational distortion of the proteinase. Serpins have been identified in animals, plants, insects, and certain viruses (1, 2). More recently, serpins have been detected in prokaryotes (3). A search of genome data bases provides evidence for Ͼ500 serpins that are grouped into 17 clades (plus Ͼ10 unclassified orphans) based upon phylogenetic relationships (4). In humans, ϳ35 serpins have been identified. These serpins are distributed among nine clades (A-I), and most are secreted and function in the circulation or extracellular spaces. These serpins regulate proteinases involved in blood coagulation, fibrinolysis, complement activation, inflammation, and extracellular matrix remodeling. In contrast, serpins belonging to clade B reside predominantly intracellularly and have been implicated in regulating apoptosis, tumor progression, and metastasis (5). However, their biological functions in terms of an intact organism have not been well defined. To date, no naturally occurring mutations with an identifiable phenotype...
SERPINB11, the last of 13 human clade B serpins to be described, gave rise to seven different isoforms. One cDNA contained a premature termination codon, two contained splice variants, and four contained full-length open reading frames punctuated by eight single nucleotide polymorphisms (SNPs). The SNPs encoded amino acid variants located within the serpin scaffold but not the reactive site loop (RSL). Although the mouse orthologue, Serpinb11, could inhibit trypsin-like peptidases, SERPINB11 showed no inhibitory activity. To determine whether the human RSL targeted a different class of peptidases or the serpin scaffold was unable to support inhibitory activity, we synthesized chimeric human and mouse proteins, in which the RSLs had been swapped. The human RSL served as a trypsin inhibitor when supported by mouse scaffold sequences. Conversely, the mouse RSL on the human scaffold showed no inhibitory activity. These findings suggested that variant residues in the SERPINB11 scaffold impaired serpin function. SDS-PAGE analysis supported this notion as RSL-cleaved SERPINB11 was unable to undergo the stressed-to-relaxed transition typical of inhibitory type serpins. Mutagenesis studies supported this hypothesis, since the reversion of amino acid sequences in helices D and I to those conserved in other clade B serpins partially restored the ability of SERPINB11 to form covalent complexes with trypsin. Taken together, these findings suggested that SER-PINB11 SNPs encoded amino acids in the scaffold that impaired RSL mobility, and HapMap data showed that the majority of genomes in different human populations harbored these noninhibitory SERPINB11 alleles. Like several other serpin superfamily members, SERPINB11 has lost inhibitory activity and may have evolved a noninhibitory function.Serpins are a superfamily of serine and cysteine peptidase inhibitors that contain ϳ1500 family members and are found in all domains of life (Eukarya, Eubacteria, and Archaea) as well as many Poxviridae (reviewed in Refs. 1-4). Unlike canonical inhibitors, inhibitory serpins employ a unique suicide substrate-like inhibitory mechanism to neutralize their target peptidases. The surface-exposed reactive site loop (RSL) 4 serves as a pseudosubstrate and binds to the active site of the peptidase. Upon cleavage of the RSL, the metastable serpin molecule undergoes a major conformational rearrangement and traps the covalently attached peptidase in a distorted, inactive form (5). A small proportion of serpins are noninhibitory and aid in diverse functions, such as hormone transport (e.g. SERPINA7/ thyroxine binding globulin) and protein folding (e.g. SER-PINH1/HSP47) (6, 7).Serpins are classified into 17 clades based on phylogenetic relationships (8). So far, 36 human serpins from nine clades (A-I) have been identified (2). The majority of serpins are plasma proteins that serve as critical regulators of important physiological processes, such as blood coagulation, fibrinolysis, and inflammation. In contrast, clade B serpins exist predominantly, but...
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