No abstract

O-acetylserine sulfhydrylase (OASS) and cystathionine β-synthase (CBS) are members of the PLP-II family, and involved in L-cysteine production. OASS produces L-cysteine via a de novo pathway while CBS participates in the reverse transsulfuration pathway. O-acetylserine-dependent CBS (OCBS) was previously identified as a new member of the PLP-II family, which are predominantly seen in bacteria. The bacterium Helicobacter pylori possess only one OASS (hp0107) gene and we showed that the protein coded by this gene actually functions as an OCBS and utilizes L-homocysteine and O-acetylserine (OAS) to produce cystathionine. HpOCBS did not show CBS activity with the substrate L-serine and required OAS exclusively. The HpOCBS structure in complex with methionine showed a closed cleft state, explaining the initial mode of substrate binding. Sequence and structural analyses showed differences between the active sites of OCBS and CBS, and explain their different substrate preferences. We identified three hydrophobic residues near the active site of OCBS, corresponding to one serine and two tyrosine residues in CBSs. Mutational studies were performed on HpOCBS and Saccharomyces cerevisiae CBS. A ScCBS double mutant (Y158F/Y226V) did not display activity with L-serine, indicating indispensability of these polar residues for selecting substrate L-serine, however, did show activity with OAS.

Mycobacterium tuberculosis (Mtb) has evolved diverse cellular processes in response to the multiple stresses it encounters within the infected host. We explored available TnSeq datasets to identify transcription factors (TFs) that are essential for Mtb survival inside the host. The analysis identified a single TF, Rv1332 (AosR), conserved across actinomycetes with a so-far uncharacterized function. AosR mitigates phagocyte-derived oxidative and nitrosative stress, thus promoting mycobacterial growth in the murine lungs and spleen. Oxidative stress induces formation of a single intrasubunit disulphide bond in AosR, which in turn facilitates AosR interaction with an extracytoplasmic-function sigma factor, SigH. This leads to the specific upregulation of the CysMdependent non-canonical cysteine biosynthesis pathway through an auxiliary intragenic stress-responsive promoter, an axis critical in detoxifying host-derived oxidative and nitrosative radicals. Failure to upregulate AosR-dependent cysteine biosynthesis during the redox stress causes differential expression of 6% of Mtb genes. Our study shows that the AosR-SigH pathway is critical for detoxifying host-derived oxidative and nitrosative radicals to enhance Mtb survival in the hostile intracellular environment.

No abstract

Human Neuroserpin (NS) a member of serine protease inhibitor (serpin) superfamily is expressed throughout the nervous system but more specifically at the later stages of neuronal cell development [1][2][3] . NS inhibits serine protease tissue-type plasminogen activator (tPA), which has been reported to play critical role in memory, synaptic plasticity and brain development 2,4,5 . Lack of NS activity or its polymerisation can directly lead to dementia and epilepsy 6,7 . NS with specific point mutations like S49P, S52R, H338R, G392E and G392R have been recognized to cause familial encephalopathy with neuroserpin inclusion bodies (FENIB) [6][7][8][9][10] . These mutations are located around the shutter region and are believed to cause opening of β -sheet A which leads to the formation of polymers via loop-sheet polymerisation mechanism 8,9,11 . Deficiency in other inhibitory serpins like α 1 -antitrypsin (AAT), α 1 -antichymotrypsin (AAC), C1-inhibitor, plasminogen activator inhibitor-1 (PAI-1) and antithrombin (AT) may result in pathological disorders like emphysema/cirrhosis, chronic obstructive bronchitis, angio-edema, bleeding disorder and thrombosis respectively [12][13][14][15][16] . These pathological disorders are collectively termed as 'serpinopathies'9 . Under normal conditions the circulatory inhibitory form of wild type (WT) serpin exists in a metastable conformation [17][18][19] . However, during its inhibitory action on serine proteases it acquires a stable conformation upon cleavage of the reactive center loop (RCL) and insertion into the β -sheet A as an additional strand 4A 20 . During this process the protease remains covalently trapped, almost irreversibly as an acyl enzyme intermediate 3 .As a result the conformational dynamics of serpin inhibition mechanism makes it prone to polymer formation. The latter is attributed to the large scale movements in RCL, helix F and strands of β -sheet A 21,22 .