Trypsin-like protease of Streptomyces exfoliatus SMF13, a potential agent in mycelial differentiation

Abstract

Streptomyces exfoliatus SMF13 sequentially produced leupeptin, leupeptininactivating enzyme (LIE) and trypsin-like protease (TLP). TLP was produced upon exhaustion of glucose. Autolysis of mycelium was accompanied by an increase in TLP activity. However, in three bld mutants isolated from 5. exfoliatus SMF13 after UV-mutagenesis, mycelium autolysis did not occur, and neither LIE nor TLP was produced, although leupeptin was produced. Production of both LIE and TLP was restored in a spontaneous Spo+ revertant of a bld mutant. In contrast, two whi mutants sequentially produced leupeptin, LIE and TLP. The molecular mass of TLP produced during morphological differentiation was estimated to be 31.8 kDa by SDS-PAGE. The N-terminal amino acid sequence was RVGGTxAAQGNFPFQQxLSM. TLP was competitively inhibited by leupeptin; the inhibition constant was 0915 pM. TLP effectively hydrolysed the mycelial protein extract of 5. exfoliatus SMF13, but the hydrolytic activity was inhibited by leupeptin. It was concluded that morphological differentiation and production of TLP are coordinately regulated, that TLP may function as an enzyme in the metabolism of mycelial proteins, and that the hydrolytic activity of TLP is regulated by autogenous leupeptin in 5. exfoliafws SMF13.

Extracellular serine-proteinases isolated from Streptomyces alboniger: Partial characterization and effect of aprotinin on cellular structure

“…are chymotrypsin-, trypsin-and subtilisin-like enzymes (Sidhu et al 1994). The optima pH and temperature values determined for these enzymes are in agreement with previous reports that described neutral to alkaline values of pH and temperatures ranging from 35°C to 75°C for proteinases purified from streptomycetes (Chandrasekaran & Dhar 1987, Kang et al 1995, Kim & Lee 1996, Yeoman & Edwards 1997. When S. alboniger was grown in the presence of aprotinin its growth was partially or completely inhibited depending on the inhibitor concentration.…”

“…These enzymes are involved in the assimilation of proteinaceous nitrogen sources, degradation of aerial mycelium and sporulation processes, as well as in antibiotic production (Ginther 1979, Kitadokoro et al 1994, Kang et al 1995, Kim & Lee 1996.…”

Purification, Characterization, and Role of Nucleases and Serine Proteases in Streptomyces Differentiation

“…Inhibitors of TLPs, such as TLCK and leupeptin, impair the formation of aerial mycelium in S. albidoflavus (33), S. exfoliatus (35), and several other species of Streptomyces (36), which suggests the widespread distribution and critical role of these proteases in bacterial development. A TLP has been proposed to play a role in the metabolism of S. exfoliatus mycelial proteins (36). No other TLP proteases have been detected in S. antibioticus throughout the developmental cycle, although a serine protease of a chymotrypsin type has also been detected in the culture medium.…”

TheStreptomycesSubtilisin Inhibitor (SSI) Gene inStreptomyces coelicolorA3(2)

“…This might also be true for several Streptomyces strains that contain a trypsin-like protease important for morphological differentiation. 10,11) In fact, the SSI gene in S. griseus is under the control of AdpA, one of the key transcriptional activators in the A-factor regulatory cascade (our unpublished data). Because several proteases are also under the control of A-factor in S. griseus, 8,9,17) study of the SSI in combination with these proteases may reveal a clue to the physiological roles of SSIs in Streptomyces.…”

“…9) In addition, in some Streptomyces species, trypsin-like proteases affect mycelial differentiation. 10,11) These observations prompted us to examine the possible involvement of SSI in aerial mycelium formation in Streptomyces as a modulator of extracellular protease activities.…”