A functional superfamily of sodium/solute symporters.

Eleven families of sodium/solute symporters are defined based on their degrees of sequence similarities, and the protein members of these families are characterized in terms of their solute and cation specificities, their sizes, their topological features, their evolutionary relationships, and their relative degrees and regions of sequence conservation. In some cases, particularly where site-specific mutagenesis analyses have provided functional information about specific proteins, multiple alignments of members of the relevant families are presented, and the degrees of conservation of the mutated residues are evaluated. Signature sequences for several of the eleven families are presented to facilitate identification of new members of these families as they become sequenced. Phylogenetic tree construction reveals the evolutionary relationships between members of each family. One of these families is shown to belong to the previously defined major facilitator superfamily. The other ten families do not show sufficient sequence similarity with each other or with other identified transport protein families to establish homology between them. This study serves to clarify structural, functional and evolutionary relationships among eleven distinct families of functionally related transport proteins.

Evolution of permease diversity and energy-coupling mechanisms with special reference to the bacterial phosphotransferase system.

Different classes of apparently unrelated permeases couple different forms of energy to solute transport. While the energy coupling mechanisms utilized by the different permease classes are clearly distinct, it is proposed, based on structural comparisons, that many of these permeases possess transmembrane, hydrophobic domains which are evolutionarily related. Carriers may have arisen from transmembrane pore-forming proteins, and the protein constituents or domains which are specifically responsible for energy coupling may have had distinct origins. Thus, complex permeases may possess mosaic structures. This suggestion is substantiated by recent findings regarding the evolutionary origins of the bacterial phosphoenolpyruvate-dependent phosphotransferase system (PTS). Mechanistic implications of this proposal are presented.

A new subfamily of bacterial ABC-type transport systems catalyzing export of drugs and carbohydrates.

Sequence comparison studies revealed that the drug resistance transporter of Streptomyces peucetius (DrrAB) and two nodulation gene products (NodIJ) of Rhizobium leguminosarum are homologous to proteins encoded by three sets of genes that comprise capsular polysaccharide export systems in gram-negative bacteria: KpsTM of Escherichia coli, BexABC of Haemophilus influenzae, and CtrDCB of Neisseria meningitidis. These five systems comprise a new subfamily within the family of ATP binding cassette (ABC)-type transporters. We have termed this subfamily the ABC-2 subfamily. For three of the systems comprising this subfamily (Drr, Nod, and Kps) only one integral membrane constituent has been identified, whereas for the other two systems (Bex and Ctr) two dis-similar integral membrane constituents have been found. This observation suggests that the transmembrane channels of ABC-2-type transporters can be formed of homo- or heterooligomers as is true of several other classes of transport systems.

A proposed link between nitrogen and carbon metabolism involving protein phosphorylation in bacteria.

We demonstrate that certain phosphoryl transfer proteins of the bacterial phosphotransferase system (PTS), the fructose- and mannitol-specific IIA proteins or domains, are homologous to a class of proteins, one of which is known to affect transcription of some of the nitrogen-regulatory sigma 54-dependent operons in Klebsiella pneumoniae. The phosphorylatable histidyl residue in the homologous PTS proteins and the consensus sequence in the vicinity of the active-site histidine are fully conserved in all members that comprise this family of proteins. A phylogenetic tree of the eight protein members of this family was constructed, and a "signature" sequence that can serve for the identification of new protein members of this family is proposed. These observations suggest that PTS-catalyzed protein phosphorylation may provide a regulatory link between carbon and nitrogen assimilation in bacteria.

A novel zinc-binding motif found in two ubiquitous deaminase families.

Two families of deaminases, one specific for cytidine, the other for deoxycytidylate, are shown to possess a novel zinc-binding motif, here designated ZBS. We have (1) identified the protein members of these 2 families, (2) carried out sequence analyses that allow specification of this zinc-binding motif, and (3) determined signature sequences that will allow identification of additional members of these families as their sequences become available.

Novel phosphotransferase system genes revealed by bacterial genome analysis: unique, putative fructose- and glucoside-specific systems.

Analyses of sequences made available through the Escherichia coli genome project in the 87.2-89.2-min and 81.5-84.5-min regions have revealed 2 putative operons encoding proteins of the phosphoenolpyruvate:sugar phosphotransferase system (PTS). The first putative operon, designated frv, includes 4 open reading frames (ORFs), ORFf147, ORFf485, ORFf356, and ORFf582, ORFf147 and ORFf485 comprise an Enzyme IIA-Enzyme IIBC pair of the PTS. The sequence similarity of ORFf485 to previously characterized fructose-specific Enzymes IIBC suggests that ORFf485 may be specific for fructose. ORFf147 encodes a protein with comparable degrees of sequence similarity to fructose and mannitol-specific Enzymes IIA as well as homologous proteins implicated in sigma 54-dependent transcriptional regulation. Unique features of this system include a detached IIA protein and the absence of a IIB domain duplication. ORFf356 and ORFf582 are functionally unidentified and nonhomologous to other ORFs in the current protein databases, but ORFf582 contains 2 N-terminal helix-turn-helix motifs, suggestive of a role in frv operon transcriptional regulation. The second putative operon, designated glv, includes 3 ORFs, ORFf455, ORFf161, and ORFf212. We suggest that ORFf455 was incorrectly assigned and should be designated ORFf368. ORFf368 and ORFf161 encode an Enzyme IIC and IIB pair of the PTS showing greatest sequence similarity to Enzymes II specific for sugars of the gluco configuration. ORFf212 encodes a protein with sequence similarity to a phospho-beta-glucosidase and an alpha-galactosidase. No putative transcriptional regulator of the glv operon was found. This operon is the first one encoding a putative PTS permease with detached Enzymes IIB and IIC and lacking an Enzyme IIA. It is suggested that both the frv and glv operons are cryptic in E. coli and that additional genes encoding novel PTS-related proteins will be revealed by bacterial genome sequence analyses.

Mammalian integral membrane receptors are homologous to facilitators and antiporters of yeast, fungi, and eubacteria.

We demonstrate that three integral membrane receptors of mammals--the ecotropic retroviral leukemia receptor (ERR), the human retroviral receptor (HRR), and the T-cell early activator (Tea)--are homologous to a family of transporters specific for amino acids, polyamines, and choline (APC), which catalyze solute uniport, solute:cation symport, or solute:solute antiport in yeast, fungi, and eubacteria. Interestingly, the ERR membrane protein was recently shown to function as a cation:amino acid cotransporter. A binary sequence similarity matrix and an evolutionary tree of the 14 members of this family, illustrating their sequence similarities and divergences, were constructed. Other proteins, including the developmentally controlled GerAII spore germination protein of Bacillus subtilis and the acetylcholine receptor of Drosophila melanogaster gave sequence comparison scores of a sufficiently large magnitude to suggest (but not to establish) a common evolutionary origin with members of the APC family. We report an extended and corrected Tea cDNA sequence and show that the mammalian Tea and ERR encoding genes are differentially expressed in tissues and cell lines. Furthermore, the two mammalian cDNA sequences hybridize with other vertebrate and yeast genomic DNAs under stringent conditions. These observations support the notion that cell surface receptor proteins in mammals are transport proteins that share a common origin with transport proteins of single-celled organisms. Thus, permeases of essential metabolites may function pathologically as viral receptors.

Sequence analyses and evolutionary relationships among the energy-coupling proteins Enzyme I and HPr of the bacterial phosphoenolpyruvate: sugar phosphotransferase system.

We have previously reported the overexpression, purification, and biochemical properties of the Bacillus subtilis Enzyme I of the phosphoenolpyruvate: sugar phosphotransferase system (PTS) (Reizer, J., et al., 1992, J. Biol. Chem. 267, 9158-9169). We now report the sequencing of the ptsI gene of B. subtilis encoding Enzyme I (570 amino acids and 63,076 Da). Putative transcriptional regulatory signals are identified, and the pts operon is shown to be subject to carbon source-dependent regulation. Multiple alignments of the B. subtilis Enzyme I with (1) six other sequenced Enzymes I of the PTS from various bacterial species, (2) phosphoenolpyruvate synthase of Escherichia coli, and (3) bacterial and plant pyruvate: phosphate dikinases (PPDKs) revealed regions of sequence similarity as well as divergence. Statistical analyses revealed that these three types of proteins comprise a homologous family, and the phylogenetic tree of the 11 sequenced protein members of this family was constructed. This tree was compared with that of the 12 sequence HPr proteins or protein domains. Antibodies raised against the B. subtilis and E. coli Enzymes I exhibited immunological cross-reactivity with each other as well as with PPDK of Bacteroides symbiosus, providing support for the evolutionary relationships of these proteins suggested from the sequence comparisons. Putative flexible linkers tethering the N-terminal and the C-terminal domains of protein members of the Enzyme I family were identified, and their potential significance with regard to Enzyme I function is discussed. The codon choice pattern of the B. subtilis and E. coli ptsI and ptsH genes was found to exhibit a bias toward optimal codons in these organisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Novel phosphotransferase-encoding genes revealed by analysis of the Escherichia coli genome: a chimeric gene encoding an Enzyme I homologue that possesses a putative sensory transduction domain.

Two genes (ptsI and ptsA) that encode homologues of the energy coupling Enzyme I of the phosphoenolpyruvate-dependent sugar-transporting phosphotransferase system (PTS) have previously been identified on the Escherichia coli chromosome. We here report the presence of a third E. coli gene, designated ptsP, that encodes an Enzyme I homologue, here designated Enzyme INtr. Enzyme INtr possesses an N-terminal domain homologous to the N-terminal domains of NifA proteins [(127 amino acids (aa)] joined via two tandem flexible linkers to the C-terminal Enzyme I-like domain (578 aa). Structural features of the putative ptsP operon, including transcriptional regulatory signals, are characterized. We suggest that Enzyme INtr functions in transcriptional regulation of nitrogen-related operons together with previously described PTS proteins encoded within the rpoN operon. It may thereby provide a link between carbon and nitrogen assimilatory pathways.

The xylose isomerase-encoding gene (xylA) of Clostridium thermosaccharolyticum: cloning, sequencing and phylogeny of XylA enzymes.

The xylose isomerase (XylA)-encoding gene (xylA) of the thermophilic anaerobic bacterium, Clostridium thermosaccharolyticum NCIB 9385, was cloned as a 4.0-kb DNA fragment by complementation of the Escherichia coli xylA mutant strain, DS941. The open reading frame of 1317 bp encoded a protein of 439 amino acids (aa), with a calculated M(r) of 50,236. The gene was preceeded by a typical clostridial Shine-Dalgarno sequence, and was expressed constitutively in the cloning host. Downstream, the clone appeared to carry a xylB gene (encoding xylulokinase) in the same orientation as xylA. Comparison of the deduced aa sequence of the C. thermosaccharolyticum XylA with 18 other XylA showed that this family of proteins was separated into two clusters, one comprising proteins from organisms with G + C-rich DNA, and the other proteins from organisms with a lower G + C composition. Within the second cluster, the XylA of C. thermosaccharolyticum was most closely related to the enzymes from C. thermosulfurogenes (Thermoanaerobacterium thermosulfurigenes) and C. thermohydrosulfuricum (93 and 84% identity, respectively). Analysis of the aligned sequences indicated two signatures (VXW[GP]GREG[YSTA]E and [LIVM]EPKPX]EQ]P) which may be useful in isolation of novel XylA.

The cellobiose permease of Escherichia coli consists of three proteins and is homologous to the lactose permease of Staphylococcus aureus.

The cellobiose (cel) operon of Escherichia coli was recently sequenced and shown to consist of five genes, celABCDF (Parker and Hall, 1990). We have shown that the CelA, CelB and CelC proteins possess amino acid sequences which are homologous to different domains of the lactose permease of Staphylococcus aureus. CelB corresponds to the integral membrane portion of the permease (IIcel) while CelC (IIIcel) and CelA (IVcel) correspond to the two cytoplasmic domains which appear to comprise the first and second phosphorylation sites in the permease, respectively. The cellobiose permease is the only one of several homologous sequenced permeases of the phosphoenolpyruvate:sugar phosphotransferase system which has its three known functional domains residing on distinct polypeptide chains.

Novel PTS proteins revealed by bacterial genome sequencing: a unique fructose-specific phosphoryl transfer protein with two HPr-like domains in Haemophilus influenzae.

The completely sequenced genome of Haemophilus influenzae has been analysed for proteins of the phosphoenolpyruvate: sugar phosphotransferase system (PTS). We show that within the fructose PTS H. influenzae possesses a novel multi-domain phosphoryl transfer protein, not previously recognized, that includes two fructose-specific HPr domains fused in tandem in a single polypeptide chain.

Sequence and evolution of the FruR protein of Salmonella typhimurium: a pleiotropic transcriptional regulatory protein possessing both activator and repressor functions which is homologous to the periplasmic ribose-binding protein.

The repressor of the fructose (fru) operon of Salmonella typhimurium (FruR) has been implicated in the transcriptional regulation of dozens of genes concerned with central metabolic pathways of carbon utilization. We here report the nucleotide sequence of the gene encoding FruR and analyse both its operator-promoter region and its deduced amino acyl sequence. The FruR protein was overexpressed and was shown to have a molecular weight of about 36 kDa in agreement with the molecular weight deduced from the gene sequence. Sequence analyses revealed that FruR is homologous to 9 distinct bacterial DNA-binding proteins, most of which recognize sugar inducers and all of which possess helix-turn-helix motifs within their N-terminal regions and exhibit sequence identity throughout most of their lengths. FruR is also homologous to the periplasmic ribose-binding protein which serves as a constituent of the ribose transport/chemoreception system. The ribose-binding protein is in turn homologous to binding proteins specific for arabinose and galactose. The periplasmic binding proteins, the structures of some of which have been elucidated in three dimensions, lack the N-terminal helix-turn-helix region, but instead possess N-terminal hydrophobic signal sequences which target them to the periplasm. A phylogenetic tree for the more closely related proteins of this superfamily was constructed, and a signature motif was identified which should facilitate future detection of additional transcriptional regulatory proteins belonging to this family.

The putative Na+/H+ antiporter (NapA) of Enterococcus hirae is homologous to the putative K+/H+ antiporter (KefC) of Escherichia coli.

Two monovalent ion porters, the putative Na+/H+ antiporter (NapA) of Enterococcus hirae and the putative K+/H+ antiporter (KefC) of Escherichia coli, are similar in sequence throughout their hydrophobic domains. These two proteins, which comprise a novel family of transporters unrelated to the previously characterized Na+/H+ exchangers of E. coli (NhaA and NhaB) are proposed to function by essentially the same mechanism.

Sequence of the fruB gene of Escherichia coli encoding the diphosphoryl transfer protein (DTP) of the phosphoenolpyruvate: sugar phosphotransferase system.

The Escherichia coli genome sequencing project in the 47 to 48 centisome region has resulted in the sequencing of the complete fructose operon (fruBKA). Due to a single base insertion, the presence of the fruB gene went unnoticed. The revised nucleotide sequence of the fruB gene, the deduced amino acid sequence of its protein product, the diphosphoryl transfer protein of the phosphoenolpyruvate: sugar phosphotransferase system, and putative transcriptional regulatory signals of the fru operon of E. coli are here presented and compared with that from Salmonella typhimurium.

Nucleotide sequence of the region between crr and cysM in Salmonella typhimurium: five novel ORFs including one encoding a putative transcriptional regulator of the phosphotransferase system.

A 4471 bp region between crr and cysM on the Salmonella typhimurium chromosome (49.5 min) has been sequenced. Five ORFs were found within this region, one of which is likely to be the putative regulatory gene, ptsJ, that corresponds in map position to a gene which when mutated allows expression of a cryptic Enzyme I of the phosphotransferase system. The deduced amino acid sequence of the encoded protein is similar to those of several open reading frames (ORFs) including ORFT2 of Rhodobacter spheroides with which it is 28% identical throughout most of its length (comparison score of 21 S.D.). PtsJ exhibits a putative, N-terminal, helix-turn-helix, DNA binding domain that is similar in sequence to those in members of the GntR family of transcriptional regulators. Analyses of the sequences of the ORFs encoded within this region are presented.

[Extremely low birth weight (less than 1000 gram) and early postnatal weight gain in preterm infants]. / Extrémen alacsony születési súlyú (< 1000 g) koraszülöttek korai postnatalis súlygyarapodása.

Improving survival of extremely low birthweight (< 1000 g) preterm infants opens the practical issues of their postnatal nutrition and growth. The authors studied nutrition and weight gain in 16 extremely low birthweight preterm infants (birthweight: 890 +/- 22 g, gestational age: 28.0 +/- 0.2 week, mean +/- SEM) during the first 12 weeks of life. Milk of the mother, or fortified pooled human milk or preterm infant formula was fed. The preterm infants approximated their birthweight by the end of the 3rd week of life (21st day: 866 +/- 29 g). Body weight expressed as per cent of birthweight was 109 +/- 2% at the end of the 4th, 176 +/- 7% at the end of the 8th and 275 +/- 6% at the end of the 12th week of life. Weight gain during the 1st to 8th postnatal weeks was compared to the mean in utero weight gain of foetuses with identical gestational age, gender and weight percentile position. Cumulative weight gain of preterm infants during the first 8 weeks of life was significantly lower than that of the theoretical controls (76 +/- 7% versus 136 +/- 2%, per cent of the initial value, preterm versus control, p < 0.0001). Additional weight gain of preterm infants was lower than that of the controls on the 1st to 5th weeks of life (g/kg/day, 1st week: -14.4 +/- 1.6 versus 16.7 +/- 0.5, p < 0.0001; 5th week: 13.3 +/- 1.2 versus 16.4 +/- 0.3, p < 0.05), there were no differences between the two groups on the 6th and 7th weeks, whereas preterm infants gained significantly more weight on the 8th week of life than the theoretical control value (18.2 +/- 0.9 versus 14.0 +/- 0.2, p < 0.001). These data indicate that the first weeks of life represent an especially important period for the improvement of the nutrition of extremely low birthweight preterm infants.