UNITED STATES

SECURITIES AND EXCHANGE COMMISSION

WASHINGTON, DC 20549

SCHEDULE 13D

Under the Securities Exchange Act of 1934

(Amendment No.     )*

Tetraphase Pharmaceuticals, Inc.

(Name of Issuer)

Common Stock, par value $0.001 per share

(Title of Class of Securities)

88165N105

(CUSIP Number)

Noubar B. Afeyan, Ph.D.

Flagship Ventures

One Memorial Drive, 7^th Floor

Cambridge, Massachusetts 02142

(617) 868-1888

with a copy to:

Sarah Rothermel, Esq.

Wilmer Cutler Pickering Hale and Dorr LLP

60 State Street

Boston, Massachusetts 02109

(617) 526-6000

(Name, Address and Telephone Number of Person Authorized to Receive Notices and Communications)

March 20, 2013

(Date of Event Which Requires Filing of This Statement)

If the filing person has previously filed a statement on Schedule 13G to report the acquisition that is the subject of this Schedule 13D, and is filing this schedule because of Rule 13d-1(e), 13d-1(f) or 13d-1(g), check the following box.  ¨

Note. Schedules filed in paper format shall include a signed original and five copies of the schedule, including all exhibits. See Rule 13d-7(b) for other parties to whom copies are to be sent.

(Continued on following pages)

(Page 1 of 17 pages)

Item 1. Security and Issuer.

This statement on Schedule 13D relates to the Reporting Persons’ (as defined in Item 2 below) beneficial ownership interest in the common stock, par value $0.001 per share (the “Common Stock”), of Tetraphase Pharmaceuticals, Inc., a Delaware Corporation (the “Issuer”). The address of the principal executive office of the Issuer is 480 Arsenal Street, Suite 110, Watertown, Massachusetts 02472. Information given in response to each item shall be deemed incorporated by reference in all other items, as applicable.

Item 2. Identity and Background.

Each of the foregoing is referred to as a “Reporting Person” and collectively as the “Reporting Persons.”

Pursuant to the provisions of General Instruction C to Schedule 13D, information in Items 2 through 6 of this Schedule 13D is provided with respect to Flagship 2004 GP, Flagship 2007 GP, Afeyan, and Kania.

Flagship Ventures

One Memorial Drive, 7^th Floor

Cambridge, Massachusetts 02142

Additional information concerning the Reporting Persons is set forth on Attachment A hereto and is incorporated by reference herein.

Item 3. Source and Amount of Funds or Other Consideration.

On August 8, 2006, the Reporting Persons acquired 2,090,000 shares of the Issuer’s Series A convertible preferred stock at a price per share of $1.00, for an aggregate purchase price of $2,090,000.00. These shares were acquired with working capital.

On June 24, 2008, the Reporting Persons acquired 2,726,086 shares of the Issuer’s Series A-2 convertible preferred stock at a price per share of approximately $1.15, for an aggregate purchase price of $3,135,000.00. These shares were acquired with working capital.

On September 11, 2009, the Reporting Persons acquired 11,422,045 shares of the Issuer’s Series B convertible preferred stock at a price per share of approximately $0.18, for an aggregate purchase price of $2,000,000.00. These shares were acquired with working capital.

On May 14, 2010, the Reporting Persons acquired 29,171,528 shares of the Issuer’s Series C convertible preferred stock at a price per share of approximately $0.26, for an aggregate purchase price of $7,500,000.00. These shares were acquired with working capital.

Following completion of the Issuer’s initial public offering on March 20, 2013 (the “IPO”), the Reporting Persons’ shares of the Issuer’s Series A convertible preferred stock, Series A-2 convertible preferred stock, Series B convertible preferred stock, and Series C convertible preferred stock converted into shares of Common Stock on a 1-for-29 basis. After the conversion of the Series A convertible preferred stock, the Series A-2 convertible preferred stock, the Series B convertible preferred stock, and the Series C convertible preferred stock, the Reporting Persons owned, in aggregate, 1,565,849 shares of the Issuer’s Common Stock.

In conjunction with the Issuer’s IPO, the Reporting Persons acquired 491,824 shares of Common Stock. These shares were acquired with working capital at the initial public offering price of $7.00 per share for an aggregate purchase price of $3,442,768.00. Following the purchase of such shares in the IPO, the Reporting Persons owned, in aggregate, 2,057,673 shares of the Issuer’s Common Stock.

Item 4. Purpose of Transaction.

The shares of Common Stock reported herein were acquired solely for investment purposes. None of the Reporting Persons have any present plans or proposals that relate to or would result in any change in the business, policies, management, structure or capitalization of the Issuer. The Reporting Persons reserve the right to acquire, or dispose of, additional securities of the Issuer in the ordinary course of their business, to the extent deemed advisable in light of their general investment and trading policies, market conditions or other factors. The Reporting Persons may engage in discussions from time to time with other stockholders of the Issuer regarding the acquisition by the Reporting Persons of shares of the Issuer’s Common Stock or other securities of the Issuer held by such stockholders.

Douglas Cole, M.D. is a member of the Board of Directors of the Issuer and is also a member of Flagship 2004 GP and Flagship 2007 GP. Mr. Cole does not own any shares of the Issuer’s Common Stock in his individual capacity and has no voting or investment control over the shares of Common Stock held by Flagship 2004 Fund or Flagship 2007 Fund. Mr. Cole disclaims beneficial ownership of the shares of Common Stock held by Flagship 2004 Fund and Flagship 2007 Fund, except to the extent of his pecuniary interest therein. Mr. Cole owns 20,000 non-qualified stock options representing the right to acquire shares of the Issuer’s Common Stock on a 1-for-1 basis. The options vest in quarterly installments over a three-year period measured from March 25, 2013.

The Reporting Persons may seek information from management and the Issuer’s Board of Directors, and may engage in further discussions with management, the Issuer’s Board of Directors, other stockholders of the Issuer and other relevant parties, concerning the business, operations, governance, management, strategy, capitalization and/or future plans of the Issuer, or in proposing one or more of the other actions described in subparagraphs (a) through (j) of this Item 4. In addition, the Reporting Persons may exercise their rights under the Investors’ Rights Agreement, as defined below.

Other than as described above and as set forth in the Investors’ Rights Agreement, as defined below, the Reporting Persons do not have any plans or proposals which would result in any of the following:

Item 5. Interest in Securities of the Issuer.

The aggregate percentage of shares of Common Stock reported beneficially owned by each person named herein has been determined in accordance with SEC rules and is based upon 19,867,967 shares of Common Stock outstanding, which is the total number of shares of Common Stock outstanding as reported in the Issuer’s final prospectus filed with the Securities and Exchange Commission on March 20, 2013 in connection with the Issuer’s IPO. The applicable SEC rules generally attribute beneficial ownership of securities to persons who possess sole or shared voting power or investment power with respect to those securities and include shares of Common Stock issuable upon the conversion or exercise of other securities that are immediately convertible or exercisable, or are convertible or exercisable within 60 days of the filing of this Report.

Please see Attachment A for additional information.

Please see Attachment A for additional information.

Please see Attachment A for additional information.

Please see Attachment A for additional information.

Please see Attachment A for additional information.

Please see Attachment A for additional information.

Item 6. Contracts, Arrangements, Understandings or Relationships with Respect to Securities of the Issuer.

Third Amended and Restated Investors’ Rights Agreement

Flagship 2004 Fund and Flagship 2007 Fund are parties to a Second Amended and Restated Registration Rights Agreement, dated as of May 14, 2010, by and among the Issuer and certain of its stockholders (the “Registration Rights Agreement”). Upon the closing of the Issuer’s IPO, holders of a total of 8,933,817 shares of Common Stock as of December 31, 2012, including for this purpose 8,828,438 shares of Common Stock issued upon conversion of the Issuer’s preferred stock upon the closing of the IPO and 54,751 shares of Common Stock issuable upon the exercise of outstanding warrants to purchase shares of the Issuer’s Series A-1 convertible preferred stock and Series C convertible preferred stock, will have the right to require the Issuer to register such shares under the Securities Act of 1933, as amended (the “Securities Act”) under specified circumstances and will have incidental registration rights as described below. After registration pursuant to these rights, these shares will become freely tradable without restriction under the Securities Act.

Certain of the Issuer’s principal stockholders who purchased shares of Common Stock in conjunction with the IPO have the registration rights described below with respect to any such purchased shares.

Demand Registration Rights

Beginning on the 180th day after March 19, 2013, subject to specified limitations set forth in the Registration Rights Agreement, at any time the holders of at least 40% of the then outstanding registrable securities, as defined in the Registration Rights Agreement, acting together, may demand in writing that the Issuer register their registrable securities under the Securities Act so long as the total amount of registrable securities registered has an aggregate proposed offering price of at least $5.0 million. The Issuer is not obligated to file a registration statement pursuant to this demand provision on more than two occasions, subject to specified exceptions.

In addition, at any time after the Issuer becomes eligible to file a registration statement on Form S-3 under the Securities Act, subject to specified limitations, the holders of at least 40% of the outstanding registrable securities may demand in writing that the Issuer register on Form S-3 the registrable securities held by them so long as the total amount of registrable securities being registered has an aggregate offering price of at least $2.0 million. The Issuer is not obligated to file a Form S-3 pursuant to this provision within six months of the effective date of any other registration statement that it may file.

Incidental Registration Rights

If, at any time after the closing of the IPO, the Issuer proposes to file a registration statement to register any of its securities under the Securities Act, either for the Issuer’s own account or for the account of any of its stockholders, other than pursuant to the demand registration rights described above and other than pursuant to a Form S-4 or Form S-8, the holders of the Issuer’s registrable securities are entitled to notice of registration and, subject to specified exceptions, the Issuer will be required to use commercially reasonable efforts to register the registrable securities then held by such holders and for which such holders request registration.

Expenses

Pursuant to the Registration Rights Agreement, the Issuer is required to pay all registration expenses, including registration and filing fees, printing expenses, fees and disbursements of its counsel and accountants, fees and expenses incurred in connection with complying with state securities or “blue sky” laws, fees of the Financial Industry Regulatory Authority, Inc., transfer taxes, fees of transfer agents and registrations, any insurance costs and reasonable fees and disbursements of one counsel representing the selling stockholders, other than any underwriting discounts and commissions, related to any demand or incidental registration. The Registration Rights Agreement contains customary cross-indemnification provisions, pursuant to which the Issuer is obligated to indemnify the selling stockholders in the event of material misstatements or omissions in the registration statement attributable to the Issuer, and the selling stockholders are obligated to indemnify the Issuer for material misstatements or omissions in the registration statement attributable to them.

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The foregoing description of the terms of the Registration Rights Agreement is intended as a summary only and is qualified in its entirety by reference to the Registration Rights Agreement, itself, which is filed as an exhibit to this Schedule 13D and incorporated by reference herein. Other than as described in this Schedule 13D, to the best of the Reporting Persons’ knowledge, there are no other contracts, arrangements, understandings or relationships (legal or otherwise) among the persons named in Item 2 and between such persons and any person with respect to any securities of the Issuer.

Item 7. Material to be Filed as Exhibits.

The following documents are filed as Exhibits to this statement:

SIGNATURES

After reasonable inquiry and to the best of my knowledge and belief, I certify that the information set forth in this statement is true, complete and correct.

Exhibit 99.1

JOINT FILING AGREEMENT

The undersigned, being duly authorized thereunder, hereby execute this agreement as an exhibit to this Schedule 13D to evidence the agreement of the below-named parties, in accordance with the rules promulgated pursuant to the Securities Exchange Act of 1934, to file this Schedule 13D jointly on behalf of each such party.

ATTACHMENT A

Afeyan and Kania are the managing members of each of Flagship 2004 GP and Flagship 2007 GP. Flagship 2004 GP is the general partner of Flagship 2004 Fund, and Flagship 2007 GP is the general partner of Flagship 2007 Fund. Each of Flagship 2004 GP, Flagship 2007 GP, Afeyan, and Kania may be deemed to possess voting and investment control over the shares of Common Stock held by Flagship 2004 Fund and Flagship 2007 Fund, and, accordingly, each of Flagship 2004 GP, Flagship 2007 GP, Afeyan, and Kania may be deemed to have indirect beneficial ownership of such shares.

Each Reporting Person disclaims beneficial ownership of the shares of the Issuer’s Common Stock described in this Schedule 13D, except to the extent of his or its pecuniary interest therein.