THE Bcl-2 family of proteins regulate programmed cell death by an unknown mechanism. Here we describe the crystal and solution structures of a Bcl-2 family member, Bcl-xL (ref. 2). The structures consist of two central, primarily hydrophobic alpha-helices, which are surrounded by amphipathic helices. A 60-residue loop connecting helices alpha1 and alpha2 was found to be flexible and non-essential for anti-apoptotic activity. The three functionally important Bcl-2 homology regions (BH1, BH2 and BH3) are in close spatial proximity and form an elongated hydrophobic cleft that may represent the binding site for other Bcl-2 family members. The arrangement of the alpha-helices in Bcl-xL is reminiscent of the membrane translocation domain of bacterial toxins, in particular diphtheria toxin and the colicins. The structural similarity may provide a clue to the mechanism of action of the Bcl-2 family of proteins.

Rate adaptation is a mechanism unspecified by the 802.11 standards, yet critical to the system performance by exploiting the multi-rate capability at the physical layer. In this paper, we conduct a systematic and experimental study on rate adaptation over 802.11 wireless networks. Our main contributions are two-fold. First, we critique five design guidelines adopted by most existing algorithms. Our study reveals that these seemingly correct guidelines can be misleading in practice, thus incur significant performance penalty in certain scenarios. The fundamental challenge is that rate adaptation must accurately estimate the channel condition despite the presence of various dynamics caused by fading, mobility and hidden terminals. Second, we design and implement a new Robust Rate Adaptation Algorithm (RRAA) that addresses the above challenge. RRAA uses short-term loss ratio to opportunistically guide its rate change decisions, and an adaptive RTS filter to prevent collision losses from triggering rate decrease. Our extensive experiments have shown that RRAA outperforms three well-known rate adaptation solutions (ARF, AARF, and SampleRate) in all tested scenarios, with throughput improvement up to 143%.

We describe the development of an expression-secretion system in Bacillus subtilis to improve the quality and quantity of the secreted foreign proteins. This system consists of a strain (WB600) deficient in six extraceilular proteases and a set of sacB-based expression vectors. With the inactivation of all six chromosomal genes encoding neutral protease A, subtilisin, extracellular protease, metalloprotease, bacillopeptidase F, and neutral protease B, WB600 showed only 0.32% of the wild-type extracellular protease activity. No residual protease activity could be detected when WB600 was cultured in the presence of 2 mM phenylmethylsulfonyl fluoride.By using TEM P-lactamase as a model, we showed that WB600 can significantly improve the stability of the secreted enzyme. To further increase the production level we constructed an expression cassette carrying sacY, a sacB-specific regulatory gene. This gene was placed under the control of a strong, constitutively expressed promoter, P43. With this cassette in the expression vector, an 18-fold enhancement in ,-lactamase production was observed. An artificial operon, P43-sacY-degQ, was also constructed. However, only a partial additive enhancement effect (24-fold enhancement) was observed. Although degQ can stimulate the production of P-lactamase in the system, its ability to increase the residual extracellular protease activity from WB600 limits its application. The use of the P43-sacY cassette and WB600 would be a better combination for producing intact foreign proteins in high yield.With the capability of secreting extracellular enzymes directly into the culture medium, Bacillus subtilis potentially can serve as an efficient expression host (4,5,9,24,25). The secreted foreign proteins usually remain in biologically active forms (20,21), and the downstream purification is greatly simplified. However, at least two major limitations hinder the wide application of B. subtilis as a production microorganism. First, B. subtilis produces and secretes high levels of extracellular proteases which degrade the secreted foreign proteins (5, 41). It is well established that B. subtilis has six extracellular proteases: neutral protease A (45), subtilisin (also known as alkaline protease) (34, 42), extracellular protease (29,37), metalloprotease (30), bacillopeptidase F (31, 43), and neutral protease B (36). The construction of a strain deficient in five proteases GP263 (30,32), has eliminated over 99% of total extracellular protease activity. However, depending on the nature of the foreign proteins to be expressed in this strain, some of them remain unstable. Second, the lack of well-regulated inducible vectors also limits the wide application of the B. subtilis system. To overcome this problem, a few inducible vectors have been developed. Most of these vectors use either the E. coli lac system (16,46) or the temperature-sensitive repressor from lambda (3) or 4105 (23) to regulate the expression of foreign genes. Some others (10,39,47,48) are based on the regulatory region of a B. ...

To study the regulatory mechanism controlling the heat-inducible expression of Bacillus subtilis groE, two regulatory elements, the A -like promoter and the inverted repeat (IR [CIRCE]) in the control region, were characterized. The groE promoter was shown to be transcribed by the major RNA polymerase under both heat shock and non-heat shock conditions. The IR was found to have two functions. (i) It ensures the fast turnover of the groE transcript, and (ii) it serves as an operator. This IR acts as a negative heat shock regulatory element, since deletion of this sequence resulted in high-level expression of groE even at 37؇C. Although this IR is present in the 5 untranslated region of the groE transcript, groE transcripts under heat shock and non-heat shock conditions showed similar in vivo half-lives of 5 min. This rapid turnover at 37؇C requires the presence of the IR. Without the IR, the groE transcript showed a longer half-life of 17 min. Increasing the distance between the groE transcription start site and the IR systematically by inserting nucleotide sequences from 5 to 21 bp in length resulted in a gradual abolition of the negative regulatory effect mediated by the IR. This effect was not due to a significant change in transcript stability or the transcription start site and is consistent with the model that this IR serves as an operator.Expression of heat shock genes in Escherichia coli is regulated by RNA polymerase containing an alternate sigma factor, 32 , encoded by rpoH (6). The transient 15-to 20-fold increase in the cellular level of 32 after heat shock plays a key role in expressing heat shock genes in this organism (7, 33). However, in Bacillus subtilis, expression of a group of heat shock genes encoding chaperonins, including GroES, GroEL, and DnaK, is regulated by a different mechanism. Two key regulatory elements are found in the regulatory regions of both the groE and dnaK operons. They are the A -like promoter and the inverted repeat (IR) sequence located between the transcription start site and the first structural gene in the operon (17,28,35). Site-directed mutagenesis of the sequence within the IR found in the B. subtilis dnaK operon results in a higher level of dnaK expression at 37ЊC, illustrating that this IR acts as a negative regulatory element (38). Furthermore, similar IRs can be found in many chaperonin operons in both gram-positive and gram-negative bacteria (20,21,25,27,29,30,34). These IRs serve as a common regulatory element in most eubacteria to control the expression of chaperones under heat shock conditions. The name ''CIRCE'' (controlling IR of chaperone expression) has been coined recently by Zuber and Schumann (38) to describe sequences in this family.Although studies (2) with in vitro transcription assays and the use of a temperature-sensitive sigA mutant support the idea that the major vegetative RNA polymerase plays a vital role in expressing these genes (1), this interpretation is complicated by the following possibilities: (i) the possible existence of trace amoun...