Catabolism of the Cholesterol Side Chain in Mycobacterium tuberculosis Is Controlled by a Redox-Sensitive Thiol Switch

Abstract

Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), is a highly successful human pathogen and has infected approximately one-third of the world’s population. Multiple drug resistant (MDR) and extensively drug resistant (XDR) TB strains and coinfection with HIV have increased the challenges of successfully treating this disease pandemic. The metabolism of host cholesterol by Mtb is an important factor for both its virulence and pathogenesis. In Mtb, the cholesterol side chain is degraded through multiple cycles of β-oxidation and FadA5 (Rv3546) catalyzes side chain thiolysis in the first two cycles. Moreover, FadA5 is important during the chronic stage of infection in a mouse model of Mtb infection. Here, we report the redox control of FadA5 catalytic activity that results from reversible disulfide bond formation between Cys59-Cys91 and Cys93-Cys377. Cys93 is the thiolytic nucleophile, and Cys377 is the general acid catalyst for cleavage of the β-keto-acyl-CoA substrate. The disulfide bond formed between the two catalytic residues Cys93 and Cys377 blocks catalysis. The formation of the disulfide bonds is accompanied by a large domain swap at the FadA5 dimer interface that serves to bring Cys93 and Cys377 in close proximity for disulfide bond formation. The catalytic activity of FadA5 has a midpoint potential of −220 mV, which is close to the Mtb mycothiol potential in the activated macrophage. The redox profile of FadA5 suggests that FadA5 is fully active when Mtb resides in the unactivated macrophage to maximize flux into cholesterol catabolism. Upon activation of the macrophage, the oxidative shift in the mycothiol potential will decrease the thiolytic activity by 50%. Thus, the FadA5 midpoint potential is poised to rapidly restrict cholesterol side chain degradation in response to oxidative stress from the host macrophage environment.

More than cholesterol catabolism: regulatory vulnerabilities in Mycobacterium tuberculosis

“…The catalytic activity of FadA5 is controlled by the reversible formation of two disulfide bonds (Cys93–Cys377 and Cys59–Cys91). The midpoint potential of FadA5 is −223 mV, which is close to the redox potential of Mtb mycothiol (−240 mV) in activated macrophages [36]. This redox matching suggests that during infection, Mtb can sense environmental changes, reduce the catalytic activity of FadA5 and thus modulate the overall production of propionyl-CoA from cholesterol.…”

Cholesterol-dependent transcriptome remodeling reveals new insight into the contribution of cholesterol to Mycobacterium tuberculosis pathogenesis

“…It is uncertain whether B 12 is continuously available in Mtb cell during infection or becomes available at some distinct stage. Nevertheless, under an unclear stimulus, which may be a change in internal redox potential and/or pH 54 , B 12 inhibits the expression of PrpR and pathways controlled by this regulator, and this change directs most of the propionate flux to virulence lipid synthesis. In addition, B 12 -unlocked MMP allows for efficient conversion of some part of carboxylated propionate into succinate, thereby bypassing down-regulated MCC and the need for anaplerosis via GC (Fig.…”

“…These genes determine entering persistence and are overexpressed in various dormancy models 80 , 81 . Importantly, the mycothiol switch controls Mtb cholesterol catabolism in response to the redox state inside the host macrophage 54 . Long-term survival of nonreplicating Mtb is ensured by the coordinated shutdown of active metabolism through the stringent response mediated by hyperphosphorylated guanosine ((p)ppGpp) 82 .…”

Degradation of Bile Acids by Soil and Water Bacteria

“…strain Chol11 [ 41 ] or R. jostii RHA1. However, the cholic acid degradation gene cluster of strain RHA1 encodes CasB [ 28 ], a homolog of the cysteine-containing thiolase FadA5, which catalyzes a thiolytic cleavage reaction of acetyl-CoA from C 5 side-chain degradation intermediates of cholesterol in M. tuberculosis H37Rv [ 92 , 93 ]. The catalytically active cysteine of typical thiolases is also conserved in CasB, indicating that acetyl-CoA is thiolytically cleaved during bile-acid degradation in strain RHA1 [ 28 ].…”