Background: Orientia tsutsugamushi (Ot) is an obligate, intracellular, gram-negative
bacterium causing scrub typhus. Some of its encoded proteins play key roles in the adhesion and internalization
of the Ot strain into host cells and are suitable resources for vaccine development and
tools for scrub typhus diagnosis. Surface cell antigen (Sca) proteins, classified as autotransporter
(AT) proteins, are one of the largest protein families involved in bacterial pathogenesis and can be
promising candidates for vaccine development. These proteins are typically large and contain inhibitory
domains; therefore, recombinant proteins without such domains have been evaluated for
this purpose. However, the expression for recombinant Sca proteins containing the AT domain,
which might largely affect their protective role against scrub typhus, has not been analyzed and optimized.
Objective: In this study, we optimized expression and purification conditions for individual Ot Sca
protein fragments [ScaA (27–1461), ScaC (257–526), ScaD (26–998), and ScaE (35–760)] harboring
the AT domain.
Methods: To this end, we subcloned sequences of codon-optimized DNA encoding Sca protein
fragments into the Escherichia coli expression vector. In addition, the expression condition for individual
Sca fragments was optimized, and the proteins were purified using one-step Ni-NTA column
method. The purified fractions were re-folded by serial dilution method, followed by BCA
quantification and densitometric analysis to estimate the yield and purity of proteins.
Results: We prepared platforms for expression of recombinant Sca protein fragments [ScaA
(27–1461), ScaC (257–526), ScaD (26–998), and ScaE (35–760)] containing an AT domain without
the signal peptide and transmembrane (TM) domain. The protein yield per liter of culture with
>70% of purity was ScaC (257–576), ScaE (35–760), ScaD (26-998), and ScaA (27-1461) in order.
Conclusion: Our results could be used to develop Sca AT-domain based vaccines and tools for
scrub typhus diagnosis with rapid and cost-effective ways.