Stuart Smith

Bio: Stuart Smith is an academic researcher from Children's Hospital Oakland Research Institute. The author has contributed to research in topics: Fatty acid synthase & Acyl carrier protein. The author has an hindex of 40, co-authored 75 publications receiving 6328 citations. Previous affiliations of Stuart Smith include University of California, San Francisco.

Conversion of a beta-ketoacyl synthase to a malonyl decarboxylase by replacement of the active-site cysteine with glutamine.

Abstract: beta-Ketoacyl synthases involved in the biosynthesis of fatty acids and polyketides exhibit extensive sequence similarity and share a common reaction mechanism, in which the carbanion participating in the condensation reaction is generated by decarboxylation of a malonyl or methylmalonyl moiety; normally, the decarboxylation step does not take place readily unless an acyl moiety is positioned on the active-site cysteine residue in readiness for the ensuing condensation reaction. Replacement of the cysteine nucleophile (Cys-161) with glutamine, in the beta-ketoacyl synthase domain of the multifunctional animal fatty acid synthase, completely inhibits the condensation reaction but increases the uncoupled rate of malonyl decarboxylation by more than 2 orders of magnitude. On the other hand, replacement with Ser, Ala, Asn, Gly, and Thr compromises the condensation reaction without having any marked effect on the decarboxylation reaction. The affinity of the beta-ketoacyl synthase for malonyl moieties, in the absence of acetyl moieties, is significantly increased in the Cys161Gln mutant compared to that in the wild type and is similar to that exhibited by the wild-type beta-ketoacyl synthase in the presence of an acetyl primer. These results, together with modeling studies of the Cys --> Gln mutant from the crystal structure of the Escherichia coli beta-ketoacyl synthase II enzyme, suggest that the side chain carbonyl group of the Gln-161 can mimic the carbonyl of the acyl moiety in the acyl-enzyme intermediate so that the mutant adopts a conformation analogous to that of the acyl-enzyme intermediate. Catalysis of the decarboxylation of malonyl-CoA requires the dimeric form of the Cys161Gln fatty acid synthase and involves prior transfer of the malonyl moiety from the CoA ester to the acyl carrier protein domain and subsequent release of the acetyl product by transfer back to a CoA acceptor. These results suggest that the role of the Cys --> Gln beta-ketoacyl synthases found in the loading domains of some modular polyketide synthases likely is to act as malonyl, or methylmalonyl, decarboxylases that provide a source of primer for the chain extension reactions catalyzed by associated modules containing fully competent beta-ketoacyl synthases.

The animal fatty acid synthase: one gene, one polypeptide, seven enzymes.

Abstract: The animal fatty acid synthase comprises two multifunctional polypeptide chains, each containing seven discrete functional domains, juxtaposed head-to-tail such that two separate centers for fatty acid assembly are formed at the subunit interface. The kinetics and specificities of the component enzymes are well adapted to ensure that, at each of the two centers, the iterative condensation of an acetyl moiety with successive malonyl moieties and complete reduction of the beta-keto intermediates normally results in the formation of palmitic acid as the major product. Nevertheless, utilization of alternative substrates and alternative chain-terminating mechanisms can extend the range of products to include branched-chain, odd carbon-numbered, and shorter chain-length fatty acids. The potential of this multifunctional form of molecular architecture for the elaboration of more complex natural products has been further exploited in microorganisms that, by the use of different fatty acid synthase "modules" that ...

Rfx6 directs islet formation and insulin production in mice and humans

Abstract: Insulin from the beta-cells of the pancreatic islets of Langerhans controls energy homeostasis in vertebrates, and its deficiency causes diabetes mellitus. During embryonic development, the transcription factor neurogenin 3 (Neurog3) initiates the differentiation of the beta-cells and other islet cell types from pancreatic endoderm, but the genetic program that subsequently completes this differentiation remains incompletely understood. Here we show that the transcription factor Rfx6 directs islet cell differentiation downstream of Neurog3. Mice lacking Rfx6 failed to generate any of the normal islet cell types except for pancreatic-polypeptide-producing cells. In human infants with a similar autosomal recessive syndrome of neonatal diabetes, genetic mapping and subsequent sequencing identified mutations in the human RFX6 gene. These studies demonstrate a unique position for Rfx6 in the hierarchy of factors that coordinate pancreatic islet development in both mice and humans. Rfx6 could prove useful in efforts to generate beta-cells for patients with diabetes.

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

Abstract: 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。

Bioorthogonal Chemistry: Fishing for Selectivity in a Sea of Functionality

Abstract: The study of biomolecules in their native environments is a challenging task because of the vast complexity of cellular systems. Technologies developed in the last few years for the selective modification of biological species in living systems have yielded new insights into cellular processes. Key to these new techniques are bioorthogonal chemical reactions, whose components must react rapidly and selectively with each other under physiological conditions in the presence of the plethora of functionality necessary to sustain life. Herein we describe the bioorthogonal chemical reactions developed to date and how they can be used to study biomolecules.

Fatty acid synthase and the lipogenic phenotype in cancer pathogenesis

Abstract: Fatty acid synthase (FASN) catalyses the synthesis of fatty acids, and this synthetic pathway is upregulated in many tumours. How might FASN and increased lipogenesis be involved in cancer, and is FASN a valid therapeutic target? There is a renewed interest in the ultimate role of fatty acid synthase (FASN) — a key lipogenic enzyme catalysing the terminal steps in the de novo biogenesis of fatty acids — in cancer pathogenesis. Tumour-associated FASN, by conferring growth and survival advantages rather than functioning as an anabolic energy-storage pathway, appears to necessarily accompany the natural history of most human cancers. A recent identification of cross-talk between FASN and well-established cancer-controlling networks begins to delineate the oncogenic nature of FASN-driven lipogenesis. FASN, a nearly-universal druggable target in many human carcinomas and their precursor lesions, offers new therapeutic opportunities for metabolically treating and preventing cancer.

Metabolic Engineering of Poly(3-Hydroxyalkanoates): From DNA to Plastic

Abstract: Poly(3-hydroxyalkanoates) (PHAs) are a class of microbially produced polyesters that have potential applications as conventional plastics, specifically thermoplastic elastomers. A wealth of biological diversity in PHA formation exists, with at least 100 different PHA constituents and at least five different dedicated PHA biosynthetic pathways. This diversity, in combination with classical microbial physiology and modern molecular biology, has now opened up this area for genetic and metabolic engineering to develop optimal PHA-producing organisms. Commercial processes for PHA production were initially developed by W. R. Grace in the 1960s and later developed by Imperial Chemical Industries, Ltd., in the United Kingdom in the 1970s and 1980s. Since the early 1990s, Metabolix Inc. and Monsanto have been the driving forces behind the commercial exploitation of PHA polymers in the United States. The gram-negative bacterium Ralstonia eutropha, formerly known as Alcaligenes eutrophus, has generally been used as the production organism of choice, and intracellular accumulation of PHA of over 90% of the cell dry weight have been reported. The advent of molecular biological techniques and a developing environmental awareness initiated a renewed scientific interest in PHAs, and the biosynthetic machinery for PHA metabolism has been studied in great detail over the last two decades. Because the structure and monomeric composition of PHAs determine the applications for each type of polymer, a variety of polymers have been synthesized by cofeeding of various substrates or by metabolic engineering of the production organism. Classical microbiology and modern molecular bacterial physiology have been brought together to decipher the intricacies of PHA metabolism both for production purposes and for the unraveling of the natural role of PHAs. This review provides an overview of the different PHA biosynthetic systems and their genetic background, followed by a detailed summation of how this natural diversity is being used to develop commercially attractive, recombinant processes for the large-scale production of PHAs.