抗原变异可使得多种致病微生物易于逃避宿主免疫应答。表达在感染红细胞表面的恶性疟原虫红细胞表面蛋白1（PfPMP1）与感染红细胞、内皮细胞、树突状细胞以及胎盘的单个或多个受体作用，在黏附及免疫逃避中起关键的作用。每个单倍体基因组var基因家族编码约60种成员，通过启动转录不同的var基因变异体为抗原变异提供了分子基础。

疟原虫var基因转换速率变化导致抗原变异[英]／Paul H, Robert P, Christodoulou Z, et al//Proc Natl Acad Sci U S A

Theoretical studies of enzymic reactions: Dielectric, electrostatic and steric stabilization of the carbonium ion in the reaction of lysozyme

Global energy and environmental problems have stimulated increased efforts towards synthesizing biofuels from renewable resources. Compared to the traditional biofuel, ethanol, higher alcohols offer advantages as gasoline substitutes because of their higher energy density and lower hygroscopicity. In addition, branched-chain alcohols have higher octane numbers compared with their straight-chain counterparts. However, these alcohols cannot be synthesized economically using native organisms. Here we present a metabolic engineering approach using Escherichia coli to produce higher alcohols including isobutanol, 1-butanol, 2-methyl-1-butanol, 3-methyl-1-butanol and 2-phenylethanol from glucose, a renewable carbon source. This strategy uses the host's highly active amino acid biosynthetic pathway and diverts its 2-keto acid intermediates for alcohol synthesis. In particular, we have achieved high-yield, high-specificity production of isobutanol from glucose. The strategy enables the exploration of biofuels beyond those naturally accumulated to high quantities in microbial fermentation.

Non-fermentative pathways for synthesis of branched-chain higher alcohols as biofuels

The phosphoenolpyruvate(PEP):carbohydrate phosphotransferase system (PTS) is found only in bacteria, where it catalyzes the transport and phosphorylation of numerous monosaccharides, disaccharides, amino sugars, polyols, and other sugar derivatives. To carry out its catalytic function in sugar transport and phosphorylation, the PTS uses PEP as an energy source and phosphoryl donor. The phosphoryl group of PEP is usually transferred via four distinct proteins (domains) to the transported sugar bound to the respective membrane component(s) (EIIC and EIID) of the PTS. The organization of the PTS as a four-step phosphoryl transfer system, in which all P derivatives exhibit similar energy (phosphorylation occurs at histidyl or cysteyl residues), is surprising, as a single protein (or domain) coupling energy transfer and sugar phosphorylation would be sufficient for PTS function. A possible explanation for the complexity of the PTS was provided by the discovery that the PTS also carries out numerous regulatory functions. Depending on their phosphorylation state, the four proteins (domains) forming the PTS phosphorylation cascade (EI, HPr, EIIA, and EIIB) can phosphorylate or interact with numerous non-PTS proteins and thereby regulate their activity. In addition, in certain bacteria, one of the PTS components (HPr) is phosphorylated by ATP at a seryl residue, which increases the complexity of PTS-mediated regulation. In this review, we try to summarize the known protein phosphorylation-related regulatory functions of the PTS. As we shall see, the PTS regulation network not only controls carbohydrate uptake and metabolism but also interferes with the utilization of nitrogen and phosphorus and the virulence of certain pathogens.

How Phosphotransferase System-Related Protein Phosphorylation Regulates Carbohydrate Metabolism in Bacteria

Publisher Summary This chapter describes many of the techniques employed to study water–macromolecules interactions, their general usefulness, points out areas of mutual support and contradiction. The chapter also presents hydration of other macromolecules and the hydration of small molecules. Four broadly based approaches to operational definitions of hydration include—(1) preferential hydration, (2) hydrodynamic hydration, (3) structural hydration, and (4) low temperature hydration. The use of high resolution X-ray and neutron diffraction data on protein crystals provides an independent approach for estimation of hydration. The chapter presents several proposals for predicting protein hydration based on the amino acid composition of the protein; however, the two main questions concerned include—whether ionic groups are more hydrated than other polar groups, and whether the amide and peptide functions are hydrated or not. Low pH, and to a much smaller degree, high pH should dehydrate proteins. Intermolecular association via ionic mechanisms should be strongly dehydrating; whereas hydrophobic interactions should not have much effect on hydration levels. Although, there is a presumption that both the enthalpy and the entropy are negative for the process of adding water molecules to isolated proteins, which are maintained in their native conformations, but there is no unequivocal evidence for this.

Hydration of Proteins and Polypeptides

The properties of acetohydroxy acid synthase (AHAS, EC 4.1.3.18) from wild-type Chlorella emersonii (var. Emersonii, CCAP-211/11n) and two spontaneous sulfometuron methyl (SMM)-resistant mutants were examined. The AHAS from both mutants was resistant to SMM and cross-resistant to imazapyr (IM) and the triazolopyrimidine sulfonanilide herbicide XRD-498 (TP). The more-SMM-resistant mutant had AHAS with altered catalytic parameters (Km, specificity), but unchanged sensitivity to the feedback inhibitors valine and leucine. The second mutant enzyme was less sensitive to the feedback inhibitors, but had otherwise unchanged kinetic parameters. Inhibition-competition experiments indicated that the three herbicides (SMM, IM, TP) bind in a mutually exclusive manner, but that valine can bind simultaneously with SMM or TP. The three herbicide classes apparently bind to closely overlapping sites. We suggest that the results with C. emersonii and other organisms can all be explained if there are separate binding sites for herbicides, feedback inhibitors and substrates.

Relationships among the herbicide and functional sites of acetohydroxy acid synthase from Chlorella emersonii

Simultaneous binding of calcium and vanadate to the Ca2+-ATPase of sarcoplasmic reticulum.

Acetohydroxyacid synthases (AHAS) and glyoxylate carboligase (GCL) catalyze decarboxylation of 2-ketoacids and condensation of the resulting hydroxyalkylThDP anions/enamines with a second ketoacid to form 2-acyl-2-hydroxyacids. AHASs prefer pyruvate by >10-fold over any other ketoacid as first substrate. Steric hindrance seems to be the major determinant of this specificity; Escherichia coli AHAS isozyme II mutant Val375Ala allows 2-ketobutyrate (C2H5COCO2−) to be a good first substrate and the mutant enzyme can thus synthesize 2-propio-2-hydroxybutyrate. An Ile residue in the equivalent position in GCL (Ile393) may play the analogous role in restricting GCL to glyoxylate (HCOCO2−) as first substrate. The specificity of AHAS for 2-ketoacids as acceptor substrates is due to an arginine residue which probably interacts with the carboxylate of the second substrate (e.g., Arg276 in AHASII). Mutants altered at this arginine can utilize aromatic aldehydes as second substrate and form chiral arylacyl carbinols, of interest as precursors for pharmaceutical syntheses. Analysis of AHAS II supports a mechanism in which carboligation occurs after rate-determining formation of hydroxyethylThDP. NMR measurements of the distribution of ThDP-bound intermediates showed that a faster rate constant for product release when the alkyl group derived from the acceptor substrate is ethyl compared to methyl plays a major role in product specificity. The crucial role of a Trp residue (Trp 464 in AHASII) in determining specificity may be due to control of a conformational change involved in product release rather than to affinity for 2-ketobutyrate. It is significant that in AHAS I, without the required Trp and with a low specificity for 2-ketobutyrate as acceptor substrate, the product release step is rapidly reversible.

Origin of the specificities of acetohydroxyacid synthases and glyoxylate carboligase

Acetohydroxyacid synthase (AHAS) activity was studied in the green unicellular alga Chlorella emersonii. This activity and its regulation was compared in the algae grown autotrophically and heterotrophically on glucose in the dark. No evidence for the existence of more than one enzyme was found. The activity in crude extracts from either heterotrophically or autotrophically grown cells showed a Km for pyruvate of 9 millimolar, a 22-fold preference for 2-ketobutyrate over pyruvate as the second substrate, 50% inhibition by 0.5 millimolar valine, and 50% inhibition by 0.3 micromolar sulfometuron methyl (SMM). Spontaneous mutants of the alga resistant to SMM were isolated, which appeared to be single gene mutants containing SMM-resistant AHAS activity. Hence, AHAS appears to be the sole direct target site of SMM in C. emersonii. The fact that the mutants had equivalent SMM resistance under auto- and heterotrophic conditions further supports the conclusion that the same enzyme functions under both physiological regimes. The addition of valine and isoleucine leads to partial relief of SMM inhibition of biomass increase, but not of SMM inhibition of cell division.

David M. Chipman

Papers

Relationships among the herbicide and functional sites of acetohydroxy acid synthase from Chlorella emersonii

Simultaneous binding of calcium and vanadate to the Ca2+-ATPase of sarcoplasmic reticulum.

Origin of the specificities of acetohydroxyacid synthases and glyoxylate carboligase

Acetohydroxy Acid Synthase Activity in Chlorella emersonii under Auto- and Heterotrophic Growth Conditions

[29] Assay of products of acetolactate synthase