A method of producing cis-4-hydroxy-L-proline, comprising the steps of: providing a L-proline cis-4-hydroxylase and L-proline; wherein the L-proline cis-4-hydroxylase is produced by expressing a DNA consisting of a nucleotide sequence encoding the amino acid sequence thereof in an expression system using a host organism and an expression vector, wherein the L-proline cis-4-hydroxylase is selected from the group consisting of the protein consisting of the amino acid sequence of SEQ ID NO: 1 or 2, a protein consisting of an amino acid sequence wherein one or several amino acids is/are deleted from, substituted in or added to the amino acid sequence of SEQ ID NO: 1 or 2, wherein the protein has L-proline cis-4-hydroxylase activity and wherein the L-proline cis-4-hydroxylase has an amino acid sequence having homology of not less than 80% with the amino acid sequence of SEQ ID NO: 1 or 2, and a protein consisting of an amino acid sequence encoded by a polynucleotide that hybridizes under stringent conditions with the nucleic acid complement of SEQ ID NO: 3 or 4, wherein the protein has L-proline cis-4-hydroxylase activity and, wherein the stringent conditions comprise hybridization overnight in a solution of 6× saline-sodium citrate and 0.2% sodium dodecyl sulfate at 65 °C, washing two times with a solution of 1 × SSC and 0.1% SDS at 65 °C for 30 minutes, and washing two times with a solution of 0.2 × SSC and 0.1% SDS at 65 °C for 30 minutes, producing cis-4-hydroxy-L-proline by a resting microbial cell reaction system or by a fermentation method, and collecting cis-4-hydroxy-L-proline. providing a L-proline cis-4-hydroxylase and L-proline; wherein the L-proline cis-4-hydroxylase is produced by expressing a DNA consisting of a nucleotide sequence encoding the amino acid sequence thereof in an expression system using a host organism and an expression vector, wherein the L-proline cis-4-hydroxylase is selected from the group consisting of the protein consisting of the amino acid sequence of SEQ ID NO: 1 or 2, a protein consisting of an amino acid sequence wherein one or several amino acids is/are deleted from, substituted in or added to the amino acid sequence of SEQ ID NO: 1 or 2, wherein the protein has L-proline cis-4-hydroxylase activity and wherein the L-proline cis-4-hydroxylase has an amino acid sequence having homology of not less than 80% with the amino acid sequence of SEQ ID NO: 1 or 2, and a protein consisting of an amino acid sequence encoded by a polynucleotide that hybridizes under stringent conditions with the nucleic acid complement of SEQ ID NO: 3 or 4, wherein the protein has L-proline cis-4-hydroxylase activity and, wherein the stringent conditions comprise hybridization overnight in a solution of 6× saline-sodium citrate and 0.2% sodium dodecyl sulfate at 65 °C, washing two times with a solution of 1 × SSC and 0.1% SDS at 65 °C for 30 minutes, and washing two times with a solution of 0.2 × SSC and 0.1% SDS at 65 °C for 30 minutes, the protein consisting of the amino acid sequence of SEQ ID NO: 1 or 2, a protein consisting of an amino acid sequence wherein one or several amino acids is/are deleted from, substituted in or added to the amino acid sequence of SEQ ID NO: 1 or 2, wherein the protein has L-proline cis-4-hydroxylase activity and wherein the L-proline cis-4-hydroxylase has an amino acid sequence having homology of not less than 80% with the amino acid sequence of SEQ ID NO: 1 or 2, and a protein consisting of an amino acid sequence encoded by a polynucleotide that hybridizes under stringent conditions with the nucleic acid complement of SEQ ID NO: 3 or 4, wherein the protein has L-proline cis-4-hydroxylase activity and, wherein the stringent conditions comprise hybridization overnight in a solution of 6× saline-sodium citrate and 0.2% sodium dodecyl sulfate at 65 °C, washing two times with a solution of 1 × SSC and 0.1% SDS at 65 °C for 30 minutes, and washing two times with a solution of 0.2 × SSC and 0.1% SDS at 65 °C for 30 minutes, producing cis-4-hydroxy-L-proline by a resting microbial cell reaction system or by a fermentation method, and collecting cis-4-hydroxy-L-proline.